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Biomedical subjects

L Duvillard

Publications and source records attributed to L Duvillard.

18 recordsLinked to original sources
PubMed2006

Inability of HDL from type 2 diabetic patients to counteract the inhibitory effect of oxidised LDL on endothelium-dependent vasorelaxation.

AIMS/HYPOTHESIS: In healthy normolipidaemic and normoglycaemic control subjects, HDL are able to reverse the inhibition of vasodilation that is induced by oxidised LDL. In type 2 diabetic patients, HDL are glycated and more triglyceride-rich than in control subjects. These alterations are likely to modify the capacity of HDL to reverse the inhibition of vasodilation induced by oxidised LDL. SUBJECTS AND METHODS: Using rabbit aorta rings, we compared the ability of HDL from 16 type 2 diabetic patients and 13 control subjects to suppress the inhibition of vasodilation that is induced by oxidised LDL. RESULTS: Oxidised LDL inhibited endothelium-dependent vasodilation (maximal relaxation [Emax] = 58.2+/-14.6 vs 99.3+/-5.2% for incubation without any lipoprotein, p < 0.0001). HDL from control subjects significantly reduced the inhibitory effect of oxidised LDL on vasodilatation (Emax = 77.6+/-12.9 vs 59.5+/-7.7%, p < 0.001), whereas HDL from type 2 diabetic patients had no effect (Emax = 52.4+/-20.4 vs 57.2+/-18.7%, NS). HDL triglyceride content was significantly higher in type 2 diabetic patients than in control subjects (5.3+/-2.2 vs 3.1+/-1.4%, p < 0.01) and was highly inversely correlated to Emax for oxidised LDL+HDL in type 2 diabetic patients (r = -0.71, p < 0.005). CONCLUSIONS/INTERPRETATION: In type 2 diabetes mellitus, the ability of HDL to counteract the inhibition of endothelium-dependent vasorelaxation induced by oxidised LDL is impaired and is inversely correlated with HDL triglyceride content. These findings suggest that HDL are less atheroprotective in type 2 diabetic patients than in control subjects.

Animals↗
PubMed2005

Normal metabolism of apolipoprotein B100-containing lipoproteins despite qualitative abnormalities in type 1 diabetic men.

AIMS/HYPOTHESIS: Type 1 diabetic subjects are at increased risk of cardiovascular disease and exhibit multiple qualitative abnormalities of apolipoprotein (apo) B100-containing lipoproteins. This stable isotope kinetic experiment was designed to study whether these abnormalities are associated with changes in the synthesis and fractional catabolic rates of VLDL-, IDL- and LDL-apoB100. METHODS: Using a bolus followed by a 16-h constant infusion of 13C-leucine, we performed a kinetic study in eight men with type 1 diabetes treated with a continuous subcutaneous insulin infusion administered by an external pump and in seven healthy men, in the fed state. RESULTS: The mean HbA1c level in the type 1 diabetic patients was 8.00+/-1.48%. Plasma triglyceride, and total, LDL and HDL cholesterol levels were similar in patients and control subjects. VLDL were less triglyceride rich in type 1 diabetic patients than in control subjects (VLDL triglyceride : apoB 6.91+/-0.81 vs 8.29+/-1.24 mmol/g, p=0.05). Conversely, the IDL and LDL of the type 1 diabetic patients contained relatively higher levels of triglycerides (IDL triglycerides : apoB 2.16+/-0.36 vs 1.57+/-0.30 mmol/g, p<0.01; LDL triglycerides : apoB 0.27+/-0.06 vs 0.16+/-0.04 mmol/g, p<0.05). The apoB100 pool size, production and fractional catabolic rates in the two groups of subjects were similar for all lipoprotein fractions. CONCLUSIONS/INTERPRETATION: Despite qualitative abnormalities, especially abnormalities of triglyceride content, the metabolism of apoB100-containing lipoproteins is not altered in type 1 diabetic men with fair glycaemic control with continuous subcutaneous insulin infusion. The high risk of atherosclerosis in these patients cannot be explained by kinetic abnormalities of apoB100-containing lipoproteins.

Adult↗
PubMed2004

Serum adiponectin and metabolic parameters in HIV-1-infected patients after substitution of nevirapine for protease inhibitors.

BACKGROUND: In the context of HIV infection and antiretroviral therapy, adiponectin concentrations have been shown to be related to lipodystrophy, metabolic alterations and HIV-protease inhibitor (PI) use. The replacement of PI by nevirapine has improved the lipid profile of patients under antiretroviral therapy. The aim of the present study was to examine whether adiponectin concentration or insulin sensitivity level correlate with the modifications of lipid parameters after the switch of PI by nevirapine. MATERIAL AND METHODS: The evolution of metabolic parameters before and after 6 months of substitution of nevirapine for protease inhibitors was evaluated in a cohort of 55 HIV-1 infected patients. Adiponectin concentration, insulin sensitivity, lipid profile, cholesterol ester transfer protein (CETP) mass concentration and triglyceride enrichment of HDL were determined before and after the replacement of PI by nevirapine. Insulin sensitivity was evaluated by the HOMA model assessment. RESULTS: Twenty-four weeks of treatment with nevirapine improved significantly the lipid profile with a significant reduction of apoB (from 0.98 to 0.92 g L(-1); P = 0.005) and triglyceride (from 2.02 to 1.66 mmol L(-1); P = 0.02). HDL cholesterol and apoA1 increased significantly (from 0.99 to 1.19 mmol L(-1); P = 0.001 and from 1.40 to 1.57 g L(-1); P < 0.001, respectively). The triglyceride enrichment of HDL significantly decreased after the replacement of PI by nevirapine (from 0.248 +/- 0.092 to 0.213 +/- 0.093; P = 0.003). At baseline, and after 24 weeks of nevirapine treatment, we observed significant correlations between adiponectin level and lipid parameters [(HDL-cholesterol (r = 0.66, P = 0.001 and r = 0.69, P = 0.001); triglycerides (r = -0.42, P = 0.002 and r = -0.57, P = 0.001), and triglyceride enrichment of HDL (r = -0.43, P = 0.005 and r = -0.53, P = 0.005)]. Twenty-four weeks of treatment with nevirapine did not significantly change adiponectin concentrations (from 984 to 1086 micro g L(-1), P = 0.22), CETP mass and insulin sensitivity. CONCLUSION: This study shows that even though a strong correlation was found between adiponectin and some metabolic parameters at baseline and after 24 weeks of treatment by nevirapine, the improvement of lipid profile observed after the replacement of PI by nevirapine was not in relation to the change of plasma adiponectin concentration. The significant decrease of triglyceride enrichment of HDL after the replacement of PI by nevirapine probably leads to a decreased catabolism of HDL lipoprotein, and consequently explains the increase of plasma HDL concentration observed in this study.

Adiponectin↗
PubMed2003

[Microbeads, nanobeads and cytometry: applications to the analysis and purification of cells and biomolecules].

Nano and microspheres are important tools in cytometry. They have been used in first to optimize fluorescent signals detected by flow cytometry and to evaluate phagocytosis. Some antigens were also detected by using nanospheres covalently coupled to antibodies. Specifically dedicated microspheres are now widely used for antigenic quantitation by flow cytometry, and magnetic nano and micropheres are very usefull for cellular and molecular purifications. To date, analytical methods based on the use of microspheres are developed to detect proteins, nucleic acids, and ions. To this end, antibodies, oligonucleotides, or chelating agents are bound to microspheres characterized by different fluorescences. The applications of these multiplexed microspheres assays allow to identify and quantify simultaneously some macromolecules and ions, but they also permit to analyze enzymatic activities and to perform polymorphism analyses. With microspheres used as reactive support, molecular analyses are therefore possible by flow cytometry. Nano and microspheres are also usefull tools for calibration in confocal microscopy as well as for micromanipulations of biomolecules and of living cells. Inovative methods based on the use of nano and microspheres are expected in the fields of biology, medicine, food industry, and environmental sciences.

Antibodies↗
PubMed2002

High-density lipoprotein apolipoprotein A-I kinetics in obese insulin resistant patients. An in vivo stable isotope study.

AIMS/HYPOTHESIS: Mechanisms responsible for the decreased high-density lipoprotein (HDL) cholesterol level associated with insulin resistance in obese patients are not clearly understood. To determine the influence of insulin resistance at an early stage on HDL metabolism, we performed a stable isotope kinetic study of apolipoprotein (apo) A-I, in five obese insulin resistant women with normal fasting triglycerides and without impaired glucose tolerance, and in five age-matched control women. METHODS: Each subject received a 16 h constant infusion of L-[1-(13)C]leucine at 0.7 mg/kg/h following a primed bolus of 0.7 mg/kg. RESULTS: ApoA-I fractional catabolic rate (FCR) was significantly increased in insulin-resistant women compared to controls (0.316+/-0.056 vs 0.210+/-0.040 per day, P<0.01), indicating a significant 50% increase of apoA-I catabolism, leading to an important reduction of plasma apoA-I residence time (3.25+/-0.59 vs 4.92+/-1.11, P<0.01). ApoA-I production rate tended to be higher in insulin resistant women than in controls (364+/-77 vs 258+/-60 mg/l/day, P=0.13), but the difference was not statistically significant. ApoA-I FCR was correlated with triglycerides during the fed state (r=0.69; P=0.026) and HDL triglycerides-esterified cholesterol ratio (r=0.73; P=0.016), suggesting that alteration of apoA-I metabolism in insulin resistance may be partly related to HDL enrichment in triglycerides. CONCLUSIONS: Our kinetic study shows that patients, at an early stage of insulin resistance (without impaired glucose tolerance nor fasting hypertriglyceridaemia), already have a significant alteration of apoA-I metabolism (increased apoA-I catabolism), which is consistent with the increased risk of atherosclerosis in this population.

Adult↗
PubMed2002

LDL-receptors expression in HIV-infected patients: relations to antiretroviral therapy, hormonal status, and presence of lipodystrophy.

BACKGROUND: Abnormalities in lipid levels and lipodystrophy (LD) have been commonly reported after commencement of highly active antiretroviral therapy (HAART). A major mechanism by which plasma low-density lipoprotein (LDL) cholesterol levels may be influenced is via the regulation of hepatic LDL receptor expression. The activity of LDL receptors is under hormonal control. Moreover, HIV infection and HAART are associated with important modifications of hormonal status. As the cause of these adverse reactions is unknown, the effects of HAART and lipodystrophy on LDL receptors were evaluated. MATERIALS AND METHODS: Thirty-nine HIV treated patients (21 with a protease inhibitor (PI) containing regimen, 18 without PI use) and 22 control subjects were tested for insulin resistance (HOMA model assessment), lipid profile, serum concentration of dehydroepiandrosterone (DHEA) and LDL-R expression. LDL-R on mononuclear cells were quantified by flow cytrometry. RESULTS: Among the 39 HIV infected patients, 14 patients had a lipodystrophy (LD). Patients with LD had significantly higher levels of triglyceride (TG) and insulin resistance compared to patients without LD. There was no significant difference in LDL-R count between patients with or without PI use. In contrast, LDL-R count was significantly lower in patients with LD compared with those without (8504 +/- 3901 vs. 13 200 +/- 4532, P = 0.001). There was no difference in LDL-R count between patients without LD and control subjects. Patients with LD had lower levels of DHEA compared to patients without LD. In HIV-infected patients, we found a significant correlation between LDL-R expression and TG (r = -0.32; P = 0.04) and LDL cholesterol (r = -0.33; P = 0.04). In contrast, we did not observe a correlation between DHEA level and LDL-R count or LDL cholesterol level. CONCLUSIONS: HIV-lipodystrophy is associated with a lower expression of LDL-R. This decreased expression of LDL-R seems independent of DHEA or insulin secretion.

Adult↗
PubMed2001

Ex vivo measurement of lipoprotein lipase-dependent very low density lipoprotein (VLDL)-triglyceride hydrolysis in human VLDL: an alternative to the postheparin assay of lipoprotein lipase activity?

The plasma lipolysis of triglyceride (TG)-rich lipoproteins is mainly due to the activity of lipoprotein lipase (LPL). Albeit important for our analysis of certain physiopathological situations, the determination of the magnitude of LPL-dependent lipolysis is not easy to perform. This essentially results from the binding of LPL to the luminal surface of vascular endothelium. The measurements of the whole putative LPL activity have been achieved after injection of heparin, a procedure that releases LPL from endothelium. However, the physiopathological relevance of this postheparin lipolysis assay (PHLA) remains questionable because it has never been demonstrated that the bulk of endothelium-bound LPL was active. It has been recently shown that a small part of LPL is associated to circulating lipoproteins in nonheparinized plasma, raising the possibility that the lipolysis mediated by this circulating LPL might reflect the overall LPL-dependent TG hydrolysis in plasma. To address this question, we developed a new lipolysis assay in which the very low density lipoprotein (VLDL)-bound LPL-dependent VLDL-TG hydrolysis (LVTH) was directly determined through the measurement of nonesterified fatty acid (NEFA) release during in vitro incubations. LVTH measurements were performed in control subjects, in type 2 diabetics, and in either heterozygous or homozygous LPL-deficient patients. In the latter group, LVTH values were extremely low. Those of heterozygous patients and of diabetics were similarly decreased by about 40% with respect to control group. Plasma TG concentrations exhibited an inverse relationship with LVTH level. In a subgroup of subjects, LVTH and PHLA were positively correlated and the inverse correlation of LVTH with plasma or VLDL-TG concentration was stronger than that obtained with PHLA. To further study the validity of this new assay, we measured LVTH in nine subjects who were studied for their catabolism of VLDL labeled with stable isotope. No relation was observed between the direct hepatic removal of VLDL and LVTH, whereas the latter was strikingly correlated with the rate of conversion of VLDL to intermediary density lipoprotein. Collective consideration of these findings strongly suggests that LVTH is a physiologically relevant index which could advantageously replace the measurements of PHLA in numerous physiopathological situations.

Adult↗
PubMed2000

Significant improvement of apolipoprotein B-containing lipoprotein metabolism by insulin treatment in patients with non-insulin-dependent diabetes mellitus.

AIMS/HYPOTHESIS: Patients with Type II (non-insulin-dependent) diabetes mellitus have multiple abnormalities in apolipoprotein B (apoB)-containing lipoprotein metabolism. These abnormalities are likely to play an important part in the development of premature atherogenesis in these patients. This stable isotope kinetic experiment was designed to study the effect of insulin therapy on apoB metabolism in poorly controlled Type II diabetic patients. METHODS: Using L-[1-13C] leucine, we studied apoB metabolism in five control subjects without insulin resistance and in six poorly controlled Type II diabetic patients before and 2 months after the introduction of insulin therapy. RESULTS: Insulin treatment induced a decrease of very low density lipoprotein apoB plasma concentration [121 +/- 42 vs 158 +/- 91 mg.l-1, p < 0.05 (control subjects: 48 +/- 20)], related to an increased catabolism of very low density lipoprotein towards intermediate density lipoprotein or low density lipoprotein [0.20 +/- 0.08 vs 0.14 +/- 0.07 pool.h-1, p < 0.05 (control subjects: 0.36 +/- 0.10)]. On the other hand, insulin treatment induced an acceleration of intermediate density lipoprotein apoB turn-over without changing its plasma concentration [77 +/- 37 vs 61 +/- 18 mg.l-1, (control subjects: 17 +/- 3)], by increasing both its production rate [22.6 +/- 9.2 vs 18.2 +/- 9.6 mg.l-1.h-1, p < 0.05 (control subjects: 18.4 +/- 3.2)] and its catabolic rate towards low density lipoprotein [0.34 +/- 0.22 vs 0.22 +/- 0.16 pool.h-1, p < 0.05 (control subjects: 1.02 +/- 0.13)]. Likewise, insulin treatment increased low density lipoprotein apoB production rate [20.2 +/- 7.4 vs 16.9 +/- 7.7 mg.l-1.h-1, p < 0.05 (control subjects: 16.9 +/- 2.3)] and restored a normal low density lipoprotein apoB fractional catabolic rate [0.022 +/- 0.004 vs 0.018 +/- 0.004 pool.h-1, p < 0.05 (control subjects: 0.025 +/- 0.004)], resulting in a constant low density lipoprotein apoB plasma concentration [965 +/- 485 vs 984 +/- 558 mg.l-1 (control subjects: 699 +/- 106)]. CONCLUSION/INTERPRETATION: Insulin treatment in Type II diabetes induces profound metabolic modifications of lipoprotein, resulting in significant decrease of the intravascular residence time of very low density lipoprotein, intermediate density lipoprotein and low density lipoprotein particles. This is likely to make these particles less harmful.

Apolipoproteins B↗
PubMed2000

Inefficiency of insulin therapy to correct apolipoprotein A-I metabolic abnormalities in non-insulin-dependent diabetes mellitus.

Non-insulin-dependent diabetes mellitus (NIDDM) is associated with low high density lipoprotein (HDL) cholesterol and apoA-I, related to an increased apoA-I fractional catabolic rate. This stable isotope kinetic experiment, using L-[1-(13)C] leucine, was designed to study the effect of insulin therapy on HDL apoA-I and A-II metabolism in poorly controlled NIDDM patients. A kinetic study was performed in five control subjects and in six NIDDM patients before and two months after the introduction of insulin therapy. ApoA-I and A-II were modelled using a monoexponential function. Insulin treatment was able to correct neither the low HDL apoA-I concentration observed in NIDDM patients (1.14+/-0.19 vs. 1.16+/-0. 12 g l(-1) (controls: 1.33+/-0.14)), nor the HDL apoA-I hypercatabolism (0.39+/-0.11 vs. 0.34+/-0.05 pool d(-1), (controls: 0.23+/-0.01, P< 0.01)). HDL apoA-I production rate was increased in NIDDM patients compared to control subjects and was not modified by insulin (0.45+/-0.12 vs. 0.39+/-0.08 g d(-1) l(-1), (controls: 0. 31+/-0.04, P< 0.05)). HDL apoA-II kinetic parameters were initially not significantly different between NIDDM patients and control subjects, and were not modified by insulin. The decreased insulin sensitivity, assessed by the insulin suppressive test, was not modified by insulin therapy in NIDDM patients. HDL apoA-I fractional catabolic rate was significantly correlated to HDL triglyceride/cholesteryl ester and triglyceride/protein ratios, which were significantly higher in NIDDM patients than in controls and were not modified by insulin therapy. The persistence of insulin resistance and of high neutral lipid exchanges between triglyceride rich lipoproteins and HDL in insulin-treated NIDDM patients probably explain the inefficiency of insulin therapy to correct HDL apoA-I metabolic abnormalities.

Adult↗
PubMed2000

HIV-1 protease inhibitors induce an increase of triglyceride level in HIV-infected men without modification of insulin sensitivity: a longitudinal study.

We investigated longitudinally the effect of protease inhibitors (PI) on insulin sensitivity, glycemia, and serum lipids in HIV-infected patients. Ninety-one consecutive patients treated with PI for at least 12 months were included in this study. Fasting glycemia, lipid profile, insulinemia, CD4 T lymphocytes, and plasma HIV-1 RNA were performed at baseline and on PI therapy. Insulin sensitivity and insulin secretion were measured by the homeostasis model assessment (HOMA MODEL) using the fasting glucose and insulin concentrations. Triglycerides (+ 0.34 mmol/l, SD = 1.07, p = 0.001) and cholesterol (+ 1.07 mmol/l, SD = 1.21, p= 0.001) significantly increased on PI therapy. Fasting glycemia, insulin sensitivity, and insulin secretion were not modified after PI therapy. PI therapy significantly increased body mass index (0.35 kg/m2, p < 0.05). Serum lipid changes correlated with changes in the CD4+ cell count. Lipodystrophy was observed in 40.6% of patients treated with PI. Our longitudinal study found that PI therapy had no major impact on fasting glycemia, insulin sensitivity, and insulin secretion. These findings are not consistent with previous cross-sectional studies, which did not include baseline measurements before PI initiation. However, we observed a similar profile of lipid changes induced by PI therapy. These results suggest that PI could be responsible for the development of hypertriglyceridemia by a mechanism independent of insulin resistance which remains to be elucidated.

Adult↗
PubMed2000

Metabolic abnormalities of apolipoprotein B-containing lipoproteins in non-insulin-dependent diabetes: a stable isotope kinetic study.

BACKGROUND: Kinetic abnormalities of apolipoprotein B (apoB)-containing lipoproteins in noninsulin-dependent diabetes mellitus (NIDDM) remain poorly understood. To get further insight into these abnormalities we performed a stable isotope kinetic experiment comparing the metabolism of apoB-containing lipoproteins in moderately severe NIDDM patients and healthy control subjects. METHODS: The study was performed in the fed state. Subjects underwent a primed infusion of 0.7 mg kg(-1) of L-[1-(13)C]leucine followed by a 16-h constant infusion of 0.7 mg kg(-1) h(-1). [13C]Leucine enrichment in apoB was measured by gas chromatography/combustion/isotope ratio mass spectrometry. RESULTS: In NIDDM patients, we observed a 3.49- and 4.52-fold increase of very-low-density lipoprotein (VLDL) and intermediate-density lipoprotein (IDL) apoB plasma concentrations, respectively (P<0.01). VLDL apoB production was increased by 41% (P<0.05) and fractional catabolic rate towards IDL and low-density lipoprotein (LDL) was decreased by 61% (P<0.05). The increased IDL apoB plasma concentration was also related to a major catabolic defect (-78%; P<0.01). For most patients, plasma LDL apoB concentration was comparable to that of controls. Nevertheless, LDL apoB metabolism was impaired in NIDDM subjects, with both a decreased LDL catabolic rate (-28%; P<0.05) and a trend towards a diminished synthesis. CONCLUSION: NIDDM is associated with multiple apoB metabolism abnormalities that are potentially atherogenic. In addition to the increased number of circulating VLDL and IDL particles, the increased residence time observed on all apoB-containing lipoproteins may promote the development of atherosclerotic lesions, by potentiating their oxidizability.

Adult↗
PubMed1998

Development of compartmental models in stable-isotope experiments: application to lipid metabolism.

Kinetic experiments are of great importance in lipid research because they further the understanding of lipid metabolism in vivo and help to explain the physiopathology of lipid disorders in humans. At present, due to species specificity, no valid animal model can efficiently replace a study in humans to explore lipid metabolism, and the use of radioactive tracers is restricted in humans. Thus, stable-isotope tracer kinetic studies have become an important component of research programs to achieve in humans a quantitative understanding of the dynamics of metabolic processes in vivo. The aim of this review is to describe the practical aspects of compartmental model development in stable-isotope experiments. The recent development of computer hardware and modeling software has dramatically facilitated the task of the modeler in his or her calculations. In the current review, we show that the model may be considered an integral component of the experimental design and that model development must obey strict rules to provide a rigorous solution. The main difficulties of model development in tracer experiments, such as experiment design, model identifiability, data expression, comparison of models, or tracer recycling, are presented with extensive references. We have paid particular attention to kinetic modeling in stable-isotope experiments because they have shown the greatest development in recent years.

Animals↗
PubMed1997

Isotope ratio mass spectrometry, compared with conventional mass spectrometry in kinetic studies at low and high enrichment levels: application to lipoprotein kinetics.

The aim of the present study was to compare the performances of gas chromatography/mass spectrometry (GC/MS) and gas chromatography/combustion/isotope ratio mass spectrometry (GC/C/IRMS) in stable isotope kinetic studies. In the analysis of cholesterol and leucine, GC/C/IRMS gave precise and linear results over a large scale of 13C enrichment (-22 to +760 delta/1000 for cholesterol, -26 to +600 delta/1000 for leucine). Compared with GC/MS, GC/C/IRMS was much more accurate and reproducible, especially at low [13C]-cholesterol enrichment (-12 delta/1000), with cholesterol samples ranging from 0.11 to 17 ng. Cholesterol ester kinetics in rabbit plasma low-density lipoproteins was studied after injection of 3 mg [3,4-(13)C]cholesterol. A smooth and regular kinetic curve was obtained with GC/C/IRMS; results were much less reproducible with GC/MS. Finally, the performances of GC/C/IRMS were demonstrated in the simultaneous kinetic study of three human plasma apolipoproteins during a primed constant infusion of 0.7 mg.kg-1.h-1 L-[l-13C]leucine. Kinetic curves were obtained in very-low-density lipoproteins and low-density lipoproteins for apolipoprotein B100, and in high-density lipoproteins for apolipoproteins AI and AIL.

Adult↗
PubMedDate not supplied

[Multicenter evaluation on different analyzers of three methods for direct HDL-cholesterol assay].

Most frequently, in routine laboratories, C-HDL is measured in the supernatant after precipitation of apolipoprotein B-containing lipoproteins by the sodium phosphotungstate/magnesium chloride reagent (PTA). This method involves precipitation, centrifugation and decantation steps which prevent full automation of the measurement and decrease the accuracy of the results. Recently, three direct assays for C-HDL including alpha-cyclodextrin sulphate (alpha-CD), polyanions/detergents (PA-D) or antibodies anti-beta-lipoproteins (AC) have been commercialized, in which all steps are fully managed by automated analyzers. These new methods have been compared to the conventional procedure (PTA), in multicenter studies among six laboratories using different analyzers. The C-HDL values measured by the alpha-CD and PA-D assays correlated well with those of the PTA method (r > 0.98), on most of the analyzers. With the AC assay, only the results obtained with the Hitachi 717 analyzer were correlated with C-HDL values of the PTA method. The linearity and specificity studies were evaluated in the laboratory A on a Kone Specific analyzer. The alpha-CD and PA-D assays were linear for C-HDL values from 0 to 5.56 mmol/l, as observed by increasing amounts of HDL2 + HDL3 or serum without lipoprotein isolated by ultracentrifugation. The specificity of these two methods was evaluated simultaneously, by adding various amounts of lipoproteins isolated by sequential ultracentrifugation. No interference was observed when adding chylomicrons up to 13.4 mmol/l of triglycerides for both methods. Inversely, increased C-HDL values were observed with added VLDL from 6 mmol/l of triglycerides for the PA-D assay and from 8 mmol/l for the alpha-CD assay. No interference was observed with added LDL up to 11.5 mmol/l of C-LDL for the alpha-CD assay and up to 6.7 mmol/l for the PA-D assay. In conclusion, the present multicenter evaluation demonstrates that the new procedures for the direct automation of C-HDL are easy and accurate and most of them correlated well with the classical precipitation method. In addition the study provides arguments for a choice between the different direct C-HDL methods.

Antibodies↗
PubMedDate not supplied

[Microspheres, nanospheres and flow cytometry: from cellular to molecular analysis].

Micro- and nanospheres are tightly associated with the development of flow cytometry. They are indispensable tools to optimize diffraction and fluorescence signals as well as for fluorescence calibration and cellular purification (magnetic micro- and nanospheres). They are also usefull to evaluate phagocytosis and to detect slightly expressed antigens. Recently, developments of microspheres-based flow cytometric assays have raised to quantify soluble analytes in biological fluids, cellular and tissue samples. The technology utilizes spectrally distinct fluorescent microspheres as a solid support for a conventional immunoassay, affinity assay or DNA hybridisation assay which is subsequently analyzed on a flow cytometer. Several multiplexed bead systems are now available facilitating the development of multiplexed assays that simultaneously measure many different analytes in few microliters of sample. Some recent applications with fluorescent microspheres coated with antibodies or oligonucleotides include cytokines and PCR products quantitation and single nucleotide polymorphism genotyping. Thus, multiplex assays using microspheres and flow cytometry technologies are exciting techniques which have the potential to contribute to the development of efficient diagnostic and research methods.

Cytological Techniques↗
PubMedDate not supplied

[Flow cytometric study of low density lipoprotein receptors: biological and clinical interest].

Low density lipoprotein (LDL) carry 60 to 80% of plasma cholesterol. These particules are mainly catabolized by LDL receptor. Thus, LDL receptor plays an important role in the regulation of plasma LDL cholesterol concentration, which is correlated to the risk of developing cardiovascular disease. The intracellular concentration of free cholesterol exerts a negative feedback on LDL receptor expression at the cell surface. This expression is also modulated by a lot of hormones, drugs and cytokines. On the other hand, mutations of LDL receptor induce familial hypercholesterolaemia, a frequent genetic disorder (1 to 500 births for the heterozygous form). Flow cytometry is a simple and fast technic, allowing to quantify LDL receptor expression at the cell surface and to study its functionality. Two different ligands can be used with flow cytometry: either fluorochrome-labelled LDL or anti-LDL receptor monoclonal antibody detected by a fluorescent secondary antibody. In immunofluorescent assays, we can now precisely calculate the number of LDL receptor at the cell surface by using calibration kits. In this article, we summarize the different regulatory factors of LDL receptor expression and present the advantages and limits of the different flow cytometry assays for LDL receptor analysis.

Flow Cytometry↗
PubMedDate not supplied

[A treatment with rosuvastatin induced a reduction of arterial pressure and a decrease of oxidative stress in spontaneously hypertensive rats].

The aim of this study was to appreciate consequences of rosuvastatin administration on hemodynamic function, vascular oxidative stress and ischemia/reperfusion disorders in normotensive and hypertensive rats. At 10 weeks of age, spontaneously hypertensive rats (SHR, n=20) and normotensive Wistar Kyoto male rats (WKY, n=20) were divided into four groups and given, either vehicle or 10 mg/kg/day of rosuvastatin by gavage for 3 weeks. Systolic blood pressure was assessed every week. At the end of these treatments, vascular NADPH oxidase activity was evaluated by chemiluminescence (lucigenin 0.5 microM). Hearts were isolated and perfused according to the Langendorff method and were subjected to 30 min of global ischemia. Reactive oxygen species (ROS) produced during reperfusion were quantified by electron spin resonance (ESR) spectroscopy using a spin probe (CP-H, 1 mM). After one week of treatment, rosuvastatin reduced the arterial pressure in SHR rats (180.3 +/- 2.1, SHR vs 169.7 +/- 2.3 mmHg, SHR+rosuvastatin; p < 0.01), without lowering plasma cholesterol levels; these effects were not observed in WKY. NADPH activity was 25% higher in control SHR rat aortas compared to control WKY, and was reduced by rosuvastatin in SHR rats. In isolated rat hearts subjected to ischemia/reperfusion sequences, there was a deterioration in functional parameters in control SHR compared to control WKY hearts. Rosuvastatin decreased post-ischemic contracture in WKY hearts by 50% (41.5 +/- 7.5, WKY control vs 18.4 +/- 4.6 mmHg, WKY+rosuvastatin; p < 0.01) and increased left ventricular developed pressure. This beneficial effect was accompanied by a decrease in ROS detected by ESR during reperfusion (312.5 +/- 45.3, WKY control; vs 219.3 +/- 22.9 AUC/mL, WKY+rosuvastatin; p < 0.05). In conclusion, these results are in accordance with the hypothesis that oxidative stress plays a crucial role in the pathogenesis of cardiovascular diseases including hypertension, and demonstrate the beneficial effects of rosuvastatin.

Animals↗