Simultaneous Rosiglitazone Release and Low-Density Lipoprotein Removal by Chondroitin Sodium Sulfate/Cyclodextrin/Poly(acrylic acid) Composite Adsorbents for Atherosclerosis Therapy.

Atherosclerosis (AS) is characterized by the accumulation of substantial low-density lipoprotein (LDL) and inflammatory response. Hemoperfusion is commonly employed for the selective removal of LDL from the body. However, conventional hemoperfusion merely focuses on LDL removal and does not address the symptom of plaque associated with AS. Based on the LDL binding properties of acrylated chondroitin sodium sulfate (CSA), acrylated beta-cyclodextrin (CD) and acrylic acid (AA), along with the anti-inflammatory property of rosiglitazone (R), the fabricated AA-CSA-CD-R microspheres could simultaneously release R and facilitate LDL removal for hemoperfusion. The AA and CSA offer electrostatic adsorption sites for LDL, while the CD provides hydrophobic adsorption sites for LDL and weak binding sites for R. According to the Sips model, the maximum static LDL adsorption capacity of AA-CSA-CD-R is determined to be 614.73 mg/g. In dynamic simulated perfusion experiments, AA-CSA-CD-R exhibits an initial cycle LDL adsorption capacity of 150.97 mg/g. The study suggests that the weakened inflammatory response favors plaque stabilization. The anti-inflammatory property of the microspheres is verified through an inflammation model, wherein the microsphere extracts are cocultured with mouse macrophages. Both qualitative analysis of iNOS\TNF-α and quantitative analysis of IL-6\TNF-α collectively demonstrate the remarkable anti-inflammatory effect of the microspheres. Therefore, the current study presents a novel blood purification treatment of eliminating pathogenic factors and introducing therapeutic factors to stabilize AS plaque.

Asymmetrical flow field-flow fractionation for improved characterization of human plasma lipoproteins.

High- and low-density lipoproteins (HDL and LDL) are attractive targets for biomarker discovery. However, ultracentrifugation (UC), the current methodology of choice for isolating HDL and LDL, is tedious, requires large sample volume, results in sample loss, and does not readily provide information on particle size. In this work, human plasma HDL and LDL are separated and collected using semi-preparative asymmetrical flow field-flow fractionation (SP-AF4) and UC. The SP-AF4 and UC separation conditions, sample throughput, and liquid chromatography/mass spectrometry (LC/MS) lipidomic results are compared. Over 600 µg of total proteins is recovered in a single SP-AF4 run, and Western blot results confirm apoA1 pure and apoB100 pure fractions, consistent with HDL and LDL, respectively. The SP-AF4 separation requires ~ 60 min per sample, thus providing a marked improvement over UC which can span hours to days. Lipidome analysis of SP-AF4-prepared HDL and LDL fractions is compared to UC-prepared HDL and LDL samples. Over 270 lipids in positive MS mode and over 140 lipids in negative MS mode are identified by both sample preparation techniques with over 98% overlap between the lipidome. Additionally, lipoprotein size distributions are determined using analytical scale AF4 coupled with multiangle light scattering (MALS) and dynamic light scattering (DLS) detectors. These developments position SP-AF4 as a sample preparation method of choice for lipoprotein biomarker characterization and identification. Graphical abstract ᅟ.

Treatment-resistant PLA2R-negative membranous nephropathy responsive to low-density lipoprotein apheresis.

Idiopathic membranous nephropathy is the most common cause of nephrotic syndrome in nondiabetic adults. The antibody most often implicated is the M-type phospholipase A2 receptor (PLA2R) antibody, found in >70% of primary membranous nephropathy cases. First-line therapy is immunosuppressive in nature, but for patients who are treatment-resistant there is a significant risk of end-stage renal disease and mortality. Hypercholesterolemia is not only a side effect of nephrotic syndrome, but also its presence may worsen renal function. A recent single-arm observational study in Japan found that low-density lipoprotein apheresis (LDL-A) was able to ameliorate nephrotic syndrome in half of patients who were resistant to medication. We present a case of treatment resistant PLA2R negative membranous nephropathy who had significant improvement following two courses of LDL-A. To our knowledge, this is the first such reported case in the United States.

Selective and Regenerable Surface Based on ß-Cyclodextrin for Low-Density Lipoprotein Adsorption.

Cyclodextrins (CDs) are a family of cyclic oligosaccharides, whose unique hydrophilic outer surface and lipophilic central cavity facilitate the formation of inclusion complexes with various biomolecules, such as cholesterol and phospholipids, via multi-interactions. Low-density lipoprotein (LDL) is the main carrier of cholesterol in bloodstream and is associated with the progression of atherosclerosis. The surface of LDL is composed of a shell of phospholipids monolayer containing most of the free unesterified cholesterol as well as the single copy of apolipoprotein B-100. To date, various LDL adsorbents have been fabricated to interact with the biomolecules on LDL surface. Owing to its elegant structure, CD is considered to be a promising choice for preparation of more economical and effective LDL-adsorbing materials. Therefore, in this study, interaction between ß-CD and LDL in solution was investigated by dynamic light scattering, circular dichroism, and ultraviolet spectroscopy. Further, a supramolecular surface based on ß-CD was simply prepared by self-assembled monolayer on gold surface. The effect of hydrogen bond and the cavity of ß-CD on the interaction between ß-CD and LDL was particularly explored by surface plasmon resonance (SPR) analysis. The SPR results showed that such ß-CD-modified surface exhibited good selectivity and could be largely regenerated by sodium dodecyl sulfate wash. This study may extend the understanding of the interaction between LDL and LDL adsorbent or the design and development of more efficient and lower-cost LDL adsorbents in the future.

Changes in serum afamin and vitamin E levels after selective LDL apheresis.

BACKGROUND: Afamin is a plasma vitamin E-binding glycoprotein partially associated with ApoA1-containing high-density lipoprotein (HDL) subfractions. In a previous study, the serum vitamin E decreased after low-density lipoprotein (LDL) apheresis, while vitamin E/cholesterol ratio increased. We aimed to study the effect of LDL apheresis on serum afamin level. METHODS: The serum level of afamin and oxidized LDL were measured by enzyme-linked immunosorbent assay in six severe heterozygous FH patients before and after their first LDL apheresis treatments and in seven healthy controls. We also investigated the changes in total cholesterol, LDL-C, HDL-C, ApoB, ApoA1, HDL subfractions, and α- and γ-tocopherol levels during the treatment. HDL subfractions were detected by an electrophoretic method on polyacrylamide gel (Lipoprint). Serum α- and γ-tocopherol levels were detected by gas chromatography-mass spectrometry. RESULTS: The first treatment sessions decreased serum afamin levels by an average of 9.4%. Total cholesterol, LDL-C, HDL-C and ApoA1 levels decreased by 52.6; 61.8; 10.5; and 14.1%, respectively. We found that α- and γ-tocopherol levels markedly decreased (by 34.1 and 32.9%, respectively), while α- tocopherol/cholesterol and γ-tocopherol/cholesterol ratios significantly increased (by 41.4 and 40.3%, respectively). Oxidized LDL levels significantly decreased. There was a shift toward the larger HDL subfractions. CONCLUSION: LDL apheresis moderately decreases the circulating levels of afamin parallel to lowering HDL-C and ApoA1 levels. Tocopherol levels decreases markedly compared to afamin levels, however, beneficial changes in vitamin E/cholesterol ratios, oxidized LDL levels and HDL subfraction distribution were detected. These additional effects of LDL apheresis may result in further cardiovascular risk reduction in FH patients.

High density lipoprotein efficiently accepts surface but not internal oxidised lipids from oxidised low density lipoprotein.

OBJECTIVE: Oxidised low density lipoprotein (oxLDL) contributes to atherosclerosis, whereas high density lipoprotein (HDL) is known to be atheroprotective due, at least in part, to its ability to remove oxidised lipids from oxLDL. The molecular details of the lipid transfer process are not fully understood. We aimed to identify major oxidised lipid species of oxLDL and investigate their transfer upon co-incubation with HDL with varying levels of oxidation. APPROACH AND RESULTS: A total of 14 major species of oxidised phosphatidylcholine and oxidised cholesteryl ester from oxLDL were identified using an untargeted mass spectrometry approach. HDL obtained from pooled plasma of normolipidemic subjects (N=5) was oxidised under mild and heavy oxidative conditions. Non-oxidised (native) HDL and oxidised HDL were co-incubated with oxLDL, re-isolated and lipidomic analysis was performed. Lipoprotein surface lipids, oxidised phosphatidylcholines and oxidised cholesterols (7-ketocholesterol and 7ß-hydroxycholesterol), but not internal oxidised cholesteryl esters, were effectively transferred to native HDL. Saturated and monounsaturated lyso-phosphatidylcholines were also transferred from the oxLDL to native HDL. These processes were attenuated when HDL was oxidised under mild and heavy oxidative conditions. The impaired capacities were accompanied by an increase in a ratio of sphingomyelin to phosphatidylcholine and a reduction in phosphatidylserine content in oxidised HDL, both of which are potentially important regulators of the oxidised lipid transfer capacity of HDL. CONCLUSIONS: Our study has revealed the differential transfer efficiency of surface and internal oxidised lipids from oxLDL and their acceptance onto HDL. These capacities were modulated when HDL was itself oxidised.

The impact of citrate concentration on adhesion of platelets and leukocytes to adsorbents in whole blood lipoprotein apheresis.

Lipoprotein apheresis is applied to deplete low density lipoprotein and other apolipoprotein B containing lipoproteins in patients with severe familial hypercholesterolemia, hypertriglyceridemia associated pancreatitis, or lipoprotein (a)-hyperlipoproteinemia. Anticoagulation of the extracorporeal circuit may influence cellular activation, as evidenced by a reduction of inflammatory parameters during regional citrate anticoagulation with acid citrate dextrose A (ACD-A) commonly used in whole blood lipid apheresis. While the citrate concentration in the extracorporeal circuit has to ensure efficient anticoagulation, citrate infusion into the patient should be limited to avoid citrate overload. We assessed the influence of citrate concentration on cellular activation during in vitro circulation of whole blood containing 2.8 mM citrate (ACD-A 1:40), 5.6 mM citrate (ACD-A 1:20), or 13 mM citrate over polyacrylate-based adsorbents for lipoprotein apheresis. We found increased platelet adhesion for anticoagulation with 2.8 mM citrate as compared to 5.6 or 13 mM citrate, as shown by cell counting and confirmed by scanning electron microscopy of adsorbent beads as well as by elevated levels of platelet activation markers and of platelet-derived microvesicles. Leukocytes showed an equivalent adhesion pattern, while red blood cells remained unaffected at all citrate concentrations. Passage of blood over two consecutive columns resulted in enhanced platelet adhesion to the second column, presumably due to upstream preactivation. In conclusion, citrate influences activation and adhesion of platelets and leukocytes in a concentration-dependent manner, and ACD-A 1:20, equivalent to a citrate concentration of 5.6 mM in whole blood, ensures minimal cellular activation during passage of whole blood over polyacrylate-based adsorbents.

Comparative evaluation of a heparin-citrate anticoagulation for LDL-apheresis in two primary apheresis systems.

BACKGROUND: As COBE Spectra has been replaced in many parts of the world, we describe a new protocol for low-density lipoprotein (LDL)-apheresis performed on familial hypercholesterolemia patients for the Spectra Optia platform. METHODS: For all procedures, after administering a bolus of heparin of 2,500 U, 10,000 U of heparin added to a 600 ml ACD-A bag was used as anticoagulant (AC). In a first phase (A), 16 apheresis procedures with COBE Spectra using an inlet:AC ratio of 25:1 were compared to 18 LDL-apheresis treatments with Spectra Optia at split Inlet:AC ratios of 16:1/18:1 or 20:1/25:1. Platelet activation and coagulation markers were assessed. In a follow-up phase (B), 20 procedures on Spectra Optia using an inlet:AC ratio of 20:1 were performed. RESULTS: Although coagulation markers and platelet activation analyzed were similar in both apheresis devices used, COBE Spectra procedures did not show any visual clumping in the sets. Visual analysis of clumping was highest in the Spectra Optia's 20:1/25:1 AC regimen (5/8 procedures). For the lowest Spectra Optia, AC regimen and during the follow-up phase reversible clump formation in the disposable set was similar (1/10 procedures). Clumping was successfully reversed in all cases by temporarily lowering the inlet:AC ratio to 18:1. Blood cell counts (WBC, Plt, Hct) were similar for both COBE Spectra and Spectra Optia procedures. Spectra Optia had a significantly higher plasma removal efficiency versus COBE Spectra (84% vs.75%, P < .05). No serious adverse events were observed. CONCLUSION: Apheresis procedures on the Spectra Optia system with low-dose heparin-citrate anticoagulation are feasible and safe.

Lowering Low-Density Lipoprotein Particles in Plasma Using Dextran Sulphate Co-Precipitates Procoagulant Extracellular Vesicles.

Plasma extracellular vesicles (EVs) are lipid membrane vesicles involved in several biological processes including coagulation. Both coagulation and lipid metabolism are strongly associated with cardiovascular events. Lowering very-low- and low-density lipoprotein ((V)LDL) particles via dextran sulphate LDL apheresis also removes coagulation proteins. It remains unknown, however, how coagulation proteins are removed in apheresis. We hypothesize that plasma EVs that contain high levels of coagulation proteins are concomitantly removed with (V)LDL particles by dextran sulphate apheresis. For this, we precipitated (V)LDL particles from human plasma with dextran sulphate and analyzed the abundance of coagulation proteins and EVs in the precipitate. Coagulation pathway proteins, as demonstrated by proteomics and a bead-based immunoassay, were over-represented in the (V)LDL precipitate. In this precipitate, both bilayer EVs and monolayer (V)LDL particles were observed by electron microscopy. Separation of EVs from (V)LDL particles using density gradient centrifugation revealed that almost all coagulation proteins were present in the EVs and not in the (V)LDL particles. These EVs also showed a strong procoagulant activity. Our study suggests that dextran sulphate used in LDL apheresis may remove procoagulant EVs concomitantly with (V)LDL particles, leading to a loss of coagulation proteins from the blood.

Human native, enzymatically modified and oxidized low density lipoproteins show different lipidomic pattern.

In the present paper we have performed comparative lipidomic analysis of two prototypic atherogenic LDL modifications, oxidized LDL and enzymatically modified LDL. Oxidization of LDL was carried out with different chemical modifications starting from the same native LDL preparations: (i) by copper oxidation leading to terminally oxidized LDL (oxLDL), (ii) by moderate oxidization with HOCl (HOCl LDL), (iii) by long term storage of LDL at 4°C to produce minimally modified LDL (mmLDL), or (iv) by 15-lipoxygenase, produced by a transfected fibroblast cell line (LipoxLDL). The enzymatic modification of LDL was performed by treatment of native LDL with trypsin and cholesteryl esterase (eLDL). Free cholesterol (FC) and cholesteryl esters (CE) represent the predominant lipid classes in all LDL preparations. In contrast to native LDL, which contains about two-thirds of total cholesterol as CE, enzymatic modification of LDL decreased the proportion of CE to about one-third. Free cholesterol and CE in oxLDL are reduced by their conversion to oxysterols. Oxidization of LDL preferentially influences the content of polyunsaturated phosphatidylcholine (PC) and polyunsaturated plasmalogen species, by reducing the total PC fraction in oxLDL. Concomitantly, a strong rise of the lysophosphatidylcholine (LPC) fraction can be found in oxLDL as compared to native LDL. This effect is less pronounced in eLDL. The mild oxidation of LDL with hypochlorite and/or lipoxygenase does not alter the content of the analyzed lipid classes and species in a significant manner. The lipidomic characterization of modified LDLs contributes to the better understanding their diverse cellular effects.

Tailor-made approach for selective isolation and elution of low-density lipoproteins by immunoaffinity sorbent on silica.

Immunoaffinity procedure was developed for isolation of low density lipoprotein (LDL) from biological samples by using silica-derived immunoaffinity sorbent. Sorbent was prepared by immobilization of monoclonal anti-apoB-100 antibody onto macroporous silica particles, using carefully optimized binding chemistry. Binding capacity of the sorbent towards LDL was determined by batch extraction experiments with solutions of isolated LDL in phosphate-buffered saline, and found to be 8 mg LDL/g. The bound LDL fraction was readily released from the sorbent by elution with ammonia at pH 11.2. The total time needed for isolation procedure was less than 1 h, with LDL recoveries being essentially quantitative for samples containing less than 0.3 mg LDL/mL. With higher concentrations, recoveries were less favorable, most probably due to irreversible adsorption caused by LDL aggreggation. However, reusability studies with isolated LDL at concentration 0.2 mg/mL indicate that the developed immunoaffinity material may be used for multiple binding-release cycles, with minor losses in binding capacity. Finally, the sorbent was successfully applied to isolation of LDL from diluted plasma. Apart from its practical implications for LDL isolation, this study provides crucial insights into issues associated with LDL-sorbent interactions, and may be useful in future efforts directed to development of lipoprotein isolation approaches.

Comparison of two low-density lipoprotein apheresis systems in patients with homozygous familial hypercholesterolemia.

Low-density lipoprotein (LDL) apheresis (LA) is a reliable method to decrease LDL-C concentrations and remains the gold standard therapy in homozygous familial hypercholesterolemia (HoFH). The objective of this study was to compare the efficacy of two LA systems [heparin-induced extracorporeal LDL precipitation (HELP) vs. dextran sulfate adsorption (DS) on the reduction of lipids, inflammatory markers, and adhesion molecules in a sample of genetically defined HoFH subjects (n = 9)]. Fasting blood samples were collected before and after LA. All subjects served as their own control and were first treated with the HELP system then with DS in this single sequence study. Compared with HELP, DS led to significantly greater reductions in total cholesterol (-63.3% vs. -59.9%; P = 0.05), LDL-C (-70.5% vs. -63.0%; P = 0.02), CRP (-75.3% vs. -48.8%; P < 0.0001), and TNF-α (-23.7% vs. +14.7%; P = 0.003). Reductions in the plasma levels of PCSK9 (-45.3% vs. -63.4%; P = 0.31), lipoprotein (a) (-70.6% vs. -65.0%; P = 0.30), E-selectin (-16.6% vs. -18.3%; P = 0.65), ICAM-1 (-4.0 vs. 5.6%; P = 0.56), and VCAM-1 (8.3% vs. -1.8%; P = 0.08) were not different between the two systems. For the same volume of filtered plasma (3,000 mL), however, HELP led to greater reductions in plasma apoB (-63.1% vs. -58.3%; P = 0.04), HDL-C (-20.6% vs. -6.5%; P = 0.003), and PCSK9 (-63.4% vs. -28.5%; P = 0.02) levels. These results suggest that both LA systems are effective in reducing plasma lipids and inflammatory markers in HoFH. Compared with HELP, greater reductions in lipid levels and inflammatory markers were achieved with DS, most likely because this method allows for a larger plasma volume to be filtered. J. Clin. Apheresis 31:359-367, 2016. © 2015 Wiley Periodicals, Inc.

[Tyramine and tryptamine as ligands for medical and biotechnological affinity sorbents].

A novel technique for preparation affinity sorbent based on tyramine and tryptamine was proposed. It was shown that tryptamine-Sepharose and tyramine-Sepharose effectively bind IgG, IgA, lipoprotein (a) (Lp(a)) and low density lipoproteins (LDL) from blood plasma. The sorption capacity is 4-9 mg of IgG, 2-4 mg IgA, 3-5:mg of Lp(a) and 5-7 mg of LDL per mL of gel. It was found that new sorbents can bind Lp(a) and IgG as themselves or in a complex of Lp(a) with IgG. The existence of this complex may indicate the presence of anti-Lp(a) autoantibodies in the blood of some patients. The advantages of new sorbents are easiness of its synthesis and stability during use and storage. In practice they can be applied for medical and biotechnological purposes where it is necessary to bind Lp(a), LDL, IgG, IgA.

Thrombocyte adhesion and release of extracellular microvesicles correlate with surface morphology of adsorbent polymers for lipid apheresis.

Whole blood lipid apheresis is clinically applied to reduce low density lipoprotein cholesterol in patients with homozygous familial hypercholesterolemia. Here, we studied the correlation between physicochemical parameters, in particular, surface roughness and blood compatibility, of two polyacrylate-based and a dextran sulfate-based polymer for lipid apheresis. The adsorbent surface roughness was assessed by atomic force microscopy. Freshly isolated human thrombocytes were circulated over adsorbent columns downscaled equivalent to clinical use to study thrombocyte adhesion and microvesicle generation. Quantification of thrombocytes and microvesicles in the flow-through of the columns revealed that both thrombocyte adhesion and microvesicle generation increased with increasing adsorbent surface roughness. Activation of thrombocytes with thrombin receptor-activating peptide-6 favored their adhesion to the adsorbents, as demonstrated by preferential depletion of CD62(+) and PAC-1(+) thrombocytes. Taken together, enhanced polymer surface roughness fostered cell adhesion and microvesicle release, underscoring the role of extracellular microvesicles as markers of cellular activation and of blood compatibility.

Recyclable heparin and chitosan conjugated magnetic nanocomposites for selective removal of low-density lipoprotein from plasma.

A new fabrication protocol is described to obtain heparin and chitosan conjugated magnetic nanocomposite as a blood purification material for removal of low-density lipoprotein (LDL) from blood plasma. The adsorbent could be easily separated with an external magnet for recyclable use since it had a magnetic core. The LDL level of plasma decreased by 67.3 % after hemoperfusion for 2 h. Moreover, the adsorbent could be recycled simply washing with NaCl solution. After eight cycles, the removal efficiency of the adsorbent was still above 50 %. The recyclable magnetic adsorbent had good blood compatibility due to the conjugation of heparin to the chitosan-coated magnetic nanocomposites. The fabricated magnetic adsorbent could be applied for LDL apheresis without side effects.

Facile fabricate sandwich-structured molecularly imprinted dopamine polymer for simultaneously specific capture of Low-density lipoprotein and eliminate "bad cholesterol".

A simplified approach for preparation of sandwich type molecularly imprinted polymers (PPDA-MIPs) is proposed for simultaneously identify Low-density lipoprotein (LDL) and dispose "bad cholesterol". Porous polydopamine nanosphere (PPDA) is applied as a matrix for immobilization of LDL, and the imprinted layer is formed by dopamine acting as a functional monomer. Since imprinted cavities exhibit shape memory effects in terms of recognizing selectivity, the PPDA-MIPs exhibit excellent selectivity toward LDL and a substantial binding capacity of 550.3 µg mg-1. Meanwhile, six adsorption/desorption cycles later, the adsorption efficiency of 83.09 % is still achieved, indicating the adequate stability and reusability of PPDA-MIPs. Additionally, over 80 % of cholesterol is recovered, indicating the completeness of "bad cholesterol" removal in LDL. Lastly, as demonstrated by gel electrophoresis, PPDA-MIPs performed satisfactory behavior for the removal of LDL from the goat serum sample.

Atorvastatin active metabolite inhibits oxidative modification of small dense low-density lipoprotein.

We tested the hypothesis that atorvastatin active metabolite (ATM), on the basis of its distinct structural features and potent antioxidant activity, preferentially inhibits lipid oxidation in human small dense low-density lipoprotein (sdLDL) and other small lipid vesicles. LDL, sdLDL, and various subfractions were isolated from human plasma by sequential ultracentrifugation, treated with ATM, atorvastatin, pravastatin, rosuvastatin, or simvastatin and were subjected to copper-induced oxidation. Lipid oxidation was measured spectrophotometrically as a function of thiobarbituric acid reactive substances formation. Similar analyses were performed in reconstituted lipid vesicles enriched in polyunsaturated fatty acids and prepared at various sizes. ATM was found to inhibit sdLDL oxidation in a dose-dependent manner. The antioxidant effects of ATM in sdLDL were 1.5 and 4.7 times greater (P < 0.001) than those observed in large buoyant LDL and very low-density lipoprotein subfractions, respectively. ATM had similar dose- and size-dependent effects in reconstituted lipid vesicles. None of these effects were reproduced by atorvastatin (parent) or any of the other statins examined in this study. These data suggest that ATM interacts with sdLDL in a specific manner that also confers preferential resistance to oxidative stress. Such interactions may reduce sdLDL atherogenicity and improve clinical outcomes in patients with cardiovascular disease.

Guaraná (Paullinia cupana Kunth) effects on LDL oxidation in elderly people: an in vitro and in vivo study.

BACKGROUND: Previous experimental investigations have suggested that guaraná (Paullinia cupana Kunth, supplied by EMBRAPA Oriental) consumption is associated with a lower prevalence of cardiovascular metabolic diseases and has positive effects on lipid metabolism, mainly related to low density lipoprotein (LDL) levels. As LDL oxidation is an important initial event in the development of atherosclerosis, we performed in vitro and in vivo studies to observe the potential effects of guaraná on LDL and serum oxidation. METHODS: The in vivo protocol was performed using blood samples from 42 healthy elderly subjects who habitually ingested guaraná (GI) or never ingested guaraná (NG). The formation of conjugated dienes (CDs) was analyzed from serum samples. The in vitro protocols were performed using LDL obtained from 3 healthy, non-fasted, normolipidemic voluntary donors who did not habitually ingest guaraná in their diets. The LDL samples were exposed to 5 different guaraná concentrations (0.05, 0.1, 0.5, 1, and 5 µg/mL). RESULTS: GI subjects demonstrated lower LDL oxidation than did NG subjects (reduction of 27%, p < 0.0014), independent of other variables. In the GI group the total polyphenols was positively associated with LDL levels. Also, guaraná demonstrated a high antioxidant activity in vitro, mainly at concentrations of 1 and 5 µg/mL, demonstrated by suppression of CDs and TBARS productions, tryptophan destruction and high TRAP activity. CONCLUSIONS: Guaraná, similar to other foods rich in caffeine and catechins such as green tea, has some effect on LDL oxidation that could partially explain the protective effects of this food in cardiometabolic diseases.

Combination treatment using percutaneous transluminal angioplasty and low-density lipoprotein apheresis in a patient with peripheral arterial disease and a history of chronic hemodialysis.

Peripheral arterial disease (PAD) is very common in dialysis patients, who tend to have diffuse calcification and severe peripheral arterial stenosis that make it difficult to treat limbs using only surgical or endovascular interventions. Better ways to treat this condition are therefore required and also follow-up studies to evaluate the effects of these treatments on the microcirculation. A 59-year-old man who had a cadaveric kidney transplant five years previously after 25 years of regular hemodialysis complained of pain at rest in his right lower limb and subsequently developed an intractable decubitus ulcer on his right fifth toe (Fontaine IV). Digital subtraction angiography revealed a severe obstruction of the right femoral artery and diffuse stenosis of the right superficial femoral artery. The patient underwent percutaneous transluminal angioplasty (PTA) and six sessions of low-density lipoprotein apheresis (LDL apheresis). At the end of these sessions his complaints were almost completely alleviated. The mean elevation in skin temperature after each session was (1.58 ± 0.99)°C [mean ± SD] over the right dorsalis pedis artery and (1.52 ± 0.88)°C at the tip of the right fifth toe. Ultrasound-measured blood flow rates in the right dorsalis pedis artery were 9.2 cm/s before PTA and 20.2 cm/s one month after PTA. Hemodialysis was resumed 3 days after the PTA due to contrast-induced nephropathy. The combination of PTA and LDL apheresis is useful for treating PAD in hemodialysis patients, with the changes in peripheral artery patency are able to be evaluated effectively by measuring skin temperature.

Measuring H(2)(18)O tracer incorporation on a QQQ-MS platform provides a rapid, transferable screening tool for relative protein synthesis.

Intracellular proteins are in a state of flux, continually being degraded into amino acids and resynthesized into new proteins. The rate of this biochemical recycling process varies across proteins and is emerging as an important consideration in drug discovery and development. Here, we developed a triple-stage quadrupole mass spectrometry assay based on product ion measurements at unit resolution and H(2)(18)O stable tracer incorporation to measure relative protein synthesis rates. As proof of concept, we selected to measure the relative in vivo synthesis rate of ApoB100, an apolipoprotein where elevated levels are associated with an increased risk of coronary heart disease, in plasma-isolated very low density lipoprotein (VLDL) and low density lipoprotein (LDL) in a mouse in vivo model. In addition, serial time points were acquired to measure the relative in vivo synthesis rate of mouse LDL ApoB100 in response to vehicle, microsomal triacylglycerol transfer protein (MTP) inhibitor, and site-1 protease inhibitor, two potential therapeutic targets to reduce plasma ApoB100 levels at 2 and 6 h post-tracer-injection. The combination of H(2)(18)O tracer with the triple quadrupole mass spectrometry platform creates an assay that is relatively quick and inexpensive to transfer across different biological model systems, serving as an ideal rapid screening tool for relative protein synthesis in response to treatment.