Niemann-Pick Type C-2 Disease: Identification by Analysis of Plasma Cholestane-3ß,5α,6ß-Triol and Further Insight into the Clinical Phenotype.

INTRODUCTION: Niemann-Pick type C disease is a rare disorder caused by impaired intracellular lipid transport due to mutations in either the NPC1 or the NPC2 gene. Ninety-five % of NPC patients show mutations in the NPC1 gene. A much smaller number of patients suffer from NPC2 disease and present respiratory failure as one of the most frequent symptoms. Several plasma oxysterols are highly elevated in NPC1 and can be used as a biomarker in the diagnosis of NPC1. METHODS: Plasma cholestane-3ß,5α,6ß-triol was evaluated as biomarker for NPC2 by GC/MS and LC-MS/MS analysis. The diagnosis was confirmed by Sanger sequencing and filipin staining. RESULTS: We report three NPC2 patients with typical respiratory problems and a detailed description of the nature of the lung disease in one of them. All patients had elevated levels of plasma cholestane-3ß,5α,6ß-triol. In two of these patients, the positive oxysterol result led to a rapid diagnosis of NPC2 by genetic analysis. The phenotype of the third patient has been described previously. In this patient a cholestane-3ß,5α,6ß-triol concentration markedly above the reference range was found. CONCLUSIONS: Measurement of plasma cholestane-3ß,5α,6ß-triol enables to discriminate between controls and NPC1 and NPC2 patients, making it a valuable biomarker for the rapid diagnosis not only for NPC1 but also for NPC2 disease.The measurement of oxysterols should be well kept in mind in the differential diagnosis of lysosomal diseases, as the elevation of oxysterols in plasma may speed up the diagnosis of NPC1 and NPC2.

Serum C-reactive protein is linked to cerebral microstructural integrity and cognitive function.

OBJECTIVE: C-reactive protein is a marker of inflammation and vascular disease. It also seems to be associated with an increased risk of dementia. To better understand potential underlying mechanisms, we assessed microstructural brain integrity and cognitive performance relative to serum levels of high-sensitivity C-reactive protein (hs-CRP). METHODS: We cross-sectionally examined 447 community-dwelling and stroke-free individuals from the Systematic Evaluation and Alteration of Risk Factors for Cognitive Health (SEARCH) Health Study (mean age 63 years, 248 female). High-field MRI was performed in 321 of these subjects. Imaging measures included fluid-attenuated inversion recovery sequences for assessment of white matter hyperintensities, automated quantification of brain parenchyma volumes, and diffusion tensor imaging for calculation of global and regional white matter integrity, quantified by fractional anisotropy (FA). Psychometric analyses covered verbal memory, word fluency, and executive functions. RESULTS: Higher levels of hs-CRP were associated with worse performance in executive function after adjustment for age, gender, education, and cardiovascular risk factors in multiple regression analysis (beta = -0.095, p = 0.02). Moreover, higher hs-CRP was related to reduced global fractional anisotropy (beta = -0.237, p < 0.001), as well as regional FA scores of the frontal lobes (beta = -0.246, p < 0.001), the corona radiata (beta = -0.222, p < 0.001), and the corpus callosum (beta = -0.141, p = 0.016), in particular the genu (beta = -0.174, p = 0.004). We did not observe a significant association of hs-CRP with measures of white matter hyperintensities or brain atrophy. CONCLUSION: These data suggest that low-grade inflammation as assessed by high-sensitivity C-reactive protein is associated with cerebral microstructural disintegration that predominantly affects frontal pathways and corresponding executive function.

Caloric restriction improves memory in elderly humans.

Animal studies suggest that diets low in calories and rich in unsaturated fatty acids (UFA) are beneficial for cognitive function in age. Here, we tested in a prospective interventional design whether the same effects can be induced in humans. Fifty healthy, normal- to overweight elderly subjects (29 females, mean age 60.5 years, mean body mass index 28 kg/m(2)) were stratified into 3 groups: (i) caloric restriction (30% reduction), (ii) relative increased intake of UFAs (20% increase, unchanged total fat), and (iii) control. Before and after 3 months of intervention, memory performance was assessed under standardized conditions. We found a significant increase in verbal memory scores after caloric restriction (mean increase 20%; P < 0.001), which was correlated with decreases in fasting plasma levels of insulin and high sensitive C-reactive protein, most pronounced in subjects with best adherence to the diet (all r values < -0.8; all P values <0.05). Levels of brain-derived neurotrophic factor remained unchanged. No significant memory changes were observed in the other 2 groups. This interventional trial demonstrates beneficial effects of caloric restriction on memory performance in healthy elderly subjects. Mechanisms underlying this improvement might include higher synaptic plasticity and stimulation of neurofacilitatory pathways in the brain because of improved insulin sensitivity and reduced inflammatory activity. Our study may help to generate novel prevention strategies to maintain cognitive functions into old age.

[The macular pigment: short- and intermediate-term changes of macular pigment optical density following supplementation with lutein and zeaxanthin and co-antioxidants. The LUNA Study]. / Kurz- und mittelfristige Anderungen der makulären Pigmentdichte infolge der Supplementation mit Lutein und Zeaxanthin sowie Koantioxidanzien. Die LUNA-Studie.

BACKGROUND: How long do the elevated concentrations of macular pigment persist after stopping supplementation with lutein and zeaxanthin? METHODS: One hundred eight (108) probands with and without age-related macular degeneration (68 female, 40 male, age 51-87 years) received a supplement containing 12 mg lutein and 1 mg zeaxanthin once per day (Ocuvite lutein) for 6 months. Analysis of macular pigment optical density (MPOD) was performed during the period of supplementation and again 3, 6, and 9 months following discontinuation of the supplement. A control group of 28 subjects received no dietary supplement. RESULTS: At baseline, the mean MPOD at 0.5 degrees was 0.50 in the supplemented group. Following supplementation, values rose, and 3 months after discontinuation of supplementation the highest levels of MPOD0.5 degrees (0.59 ODU) were detected (increase of +0.1 ODU, (p<0.001). Six months after supplement discontinuation, a slight decrease of mean MPOD0.5 degrees appeared (to 0.54 ODU), followed again by a slight increase 3 months later (to 0.57 ODU). An increment of MPOD0.5 degrees in the control group was not significant (0.03 ODU, p=0.15). DISCUSSION: Supplementation of lutein and zeaxanthin leads to an increase of MPOD0.5 degrees . This effect outlasts the duration of intake, and 9 months after supplementation was stopped, the mean MPOD0.5 degrees was still elevated compared with baseline levels. A longer follow-up subsequent to stopping supplementation might clarify whether the values decrease over time or whether a plateau of elevated MPOD levels is reached. High doses of lutein and zeaxanthin seem to be necessary to increase macular pigment density in the retina; afterwards, the amount of carotenoids needed to maintain high concentrations seems to be covered by daily food.

Exercise-induced redistribution of T lymphocytes is regulated by adrenergic mechanisms.

Acute exercise is known for causing considerable changes in leukocyte counts and function. In this paper we report that differentiated changes in T-lymphocyte distribution occur in lymphoid and non-lymphoid organs depending on the type and the intensity of exercise. Using fluorescent cell tracking we observed a release of T-cells from the spleen while lung, bone marrow and Peyer's patches served as target organs. The number of T-cells in the blood rose after intensive running while lymphopenia occurred after swimming exercise. Changes in number of labelled T-cells were neither found in the lymph nodes nor in the thymus regardless of exercise protocol. Following an alpha- or beta-blockade, the exercise-induced release of T-cells from the spleen and the accumulation of T-cells in the lung were inhibited while the enhancement of T-cells in the Peyer's patches was not affected. The administration of epinephrine partially mimicked the effects of exercise and resulted in a release of T-cells from both, the spleen and the liver, as well as in an increase of circulating blood T-cells. In conclusion, exercise induces a substantial re-distribution of T-cells within lymphoid and non-lymphoid organs. The migrating properties of T-cells could be partially explained by adrenergic mechanisms associated with exercise while the involvement of certain homing receptors remains to be shown. Our results suggest that the accumulation of T-cells in both, lung and Peyer's patches, may enhance the immune vigilance in these compartments which serve as the body's major defence barriers.

Impaired vitamin B12 metabolic status in healthcare workers occupationally exposed to nitrous oxide.

BACKGROUND: Previous studies demonstrated inactivation of vitamin B12 by nitrous oxide (N(2)O). The intraoperative exposure to N(2)O was shown to induce megaloblastic anaemia and myelopathy in subjects with subclinical vitamin B12 deficiency. In contrast, no data concerning the influence of occupational exposure to N(2)O on vitamin B12 metabolic status are available to date. In the present study, the vitamin B12 status in operating theatre personnel was assessed in relation to the extent of exposure. METHODS: Ninety-five operating theatre nurses with the history of exposure to N(2)O and 90 unexposed counterparts were examined. Vitamin B12 and folic acid were measured by immunoassay. Total homocysteine (tHcy), an indicator of impaired vitamin B12 metabolism, was determined by high performance liquid chromatography. N(2)O concentration was monitored by adsorption gas chromatography and mass spectrometry. RESULTS: No significant differences were found between both groups with respect to haematological parameters and folic acid. However, subjects exposed to N(2)O presented with lower vitamin B12 [372.8 (12.1) vs 436.8 (13.2) pmol litre(-1), P<0.001] and higher tHcy [11.2 (0.5) vs 8.9 (0.5) micromol litre(-1), P=0.006]. The changes in vitamin B12 status were aggravated in subjects exposed to N(2)O in concentrations substantially exceeding occupational exposure limit (180 mg m(-3)) [vitamin B12: 341.9 (17.7) vs 436.8 (13.2) pmol litre(-1), P=0.006; tHcy: 12.9 (0.7) vs 8.9 (0.5) micromol litre(-1), P=0.047]. CONCLUSIONS: Exposure to N(2)O in healthcare workers is associated with alterations of vitamin B12 metabolic status, the extent of which depends on the level of exposure.

Influence of three rapeseed oil-rich diets, fortified with alpha-linolenic acid, eicosapentaenoic acid or docosahexaenoic acid on the composition and oxidizability of low-density lipoproteins: results of a controlled study in healthy volunteers.

OBJECTIVE: To compare the individual effects of dietary alpha-linolenic acid (ALA), eicosapentaenoic acid (EPA) or docosahexaenoic acid (DHA) on low-density lipoprotein (LDL) fatty acid composition, ex vivo LDL oxidizability and tocopherol requirement. DESIGN, SETTING AND SUBJECTS: A randomized strictly controlled dietary study with three dietary groups and a parallel design, consisting of two consecutive periods. Sixty-one healthy young volunteers, students at a nearby college, were included. Forty-eight subjects (13 males, 35 females) completed the study. INTERVENTIONS: Subjects received a 2-week wash-in diet rich in monounsaturated fatty acids (21% energy) followed by experimental diets enriched with about 1% of energy of ALA, EPA or DHA for 3 weeks. The omega-3 (n-3) fatty acids were provided with special rapeseed oils and margarines. The wash-in diet and the experimental diets were identical, apart from the n-3 fatty acid composition and the tocopherol content, which was adjusted to the content of dienoic acid equivalents. RESULTS: Ex vivo oxidative susceptibility of LDL was highest after the DHA diet, indicated by a decrease in lag time (-16%, P<0.001) and an increase in the maximum amount of conjugated dienes (+7%, P<0.001). The EPA diet decreased the lag time (-16%, P<0.001) and the propagation rate (-12%, P<0.01). Tocopherol concentrations in LDL decreased in the ALA group (-13.5%, P<0.05) and DHA group (-7.3%, P<0.05). Plasma contents of tocopherol equivalents significantly decreased in all three experimental groups (ALA group: -5.0%, EPA group: -5.7%, DHA group: -12.8%). The content of the three n-3 polyunsaturated fatty acid differently increased in the LDL: on the ALA diet, the ALA content increased by 89% (P<0.001), on the EPA diet the EPA content increased by 809% (P<0.001) and on the DHA diet, the DHA content increased by 200% (P<0.001). In addition, the EPA content also enhanced (without dietary intake) in the ALA group (+35%, P<0.01) and in the DHA group (+284%, P<0.001). CONCLUSIONS: Dietary intake of ALA, EPA or DHA led to a significant enrichment of the respective fatty acid in the LDL particles, with dietary EPA preferentially incorporated. In the context of a monounsaturated fatty acid-rich diet, ALA enrichment did not enhance LDL oxidizability, whereas the effects of EPA and DHA on ex vivo LDL oxidation were inconsistent, possibly in part due to further changes in LDL fatty acid composition.

A randomized controlled trial comparing hydration therapy to additional hemodialysis or N-acetylcysteine for the prevention of contrast medium-induced nephropathy: the Dialysis-versus-Diuresis (DVD) Trial.

Contrast medium-induced nephropathy (CIN) is a serious complication with increasing frequency and an unfavorable prognosis. Previous analyses of surrogate parameters have suggested beneficial effects of hemodialysis that are assessed in this randomized clinical trial. We performed a prospective single-center trial in 424 consecutive patients with serum creatinine concentrations between 1.3- 3.5 mg/dl who underwent elective coronary angiography. Patients were randomized to one of three treatment strategies with all patients receiving pre- and postprocedural hydration: One group received no additional therapy, patients in the second group were hemodialyzed once, and the third group received oral N-acetylcysteine. The frequency of CIN (defined as an increase in serum creatinine>or=0.5 mg/dl) from 48 to 72 h after catheterization was 6.1% in the hydration-only group, 15.9% with hemodialysis treatment, and 5.3% in the N-ACC group (intention-to-treat analysis; P=0.008). There were no differences between the treatment groups with regard to increased (>or=0.5 mg/dl) serum creatinine concentrations after 30-60 days (4.8%, 5.1%, and 3.1%, respectively; P=0.700). Analyses of long-term follow-up (range 63 to 1316 days) by Cox regressions models of the study groups found quite similar survival rates (P=0.500). In contrast to other (retrospective) studies, long-term survival of patients with vs those without CIN within 72 h was not different, but patients who still had elevated creatinine concentrations at 30-60 days suffered from a markedly higher 2-year mortality (46% vs 17%, P=0.002). In conclusion, hemodialysis in addition to hydration therapy for the prevention of CIN provided no evidence for any outcome benefit but evidence for probable harm. Increased creatinine concentrations at 30-60 days, but not within 72 h, were associated with markedly reduced long-term survival.

The response of the novel pro-inflammatory molecules S100A8/A9 to exercise.

Exercise shares many similarities with the acute phase response of inflammatory diseases. Recently, elevated serum levels of the novel pro-inflammatory molecules of the S100 protein family, S100A8 and S100A9, have been associated with various inflammatory diseases. The present study was conducted to assess their potential roles as inflammatory markers in monitoring the exercise-induced immune response. Seventeen male subjects of different training status performed a marathon run. Furthermore 13 subjects (10 male, 3 female) performed three different treadmill tests: strenuous (STE), moderate (MTE), and downhill (DTE). S100A8/A9 complexes were measured by ELISA, while white blood cell count (WBC) and C-reactive protein (CRP) were used as markers of the inflammatory response. Serum creatine kinase (CK) concentration was determined as a marker for muscle damage. After marathon S100A8/A9 increased dramatically during the early post-exercise period and returned to resting levels one day after the run. A similar pattern was found for WBC, while CK and CRP reached their maximum on the day after the run. Moreover, S100A8/A9 release was higher in the subgroup of well-trained athletes. The kinetic of the S100A8/A9 release after the treadmill tests depended on exercise intensity and was prolonged after eccentric exercise. In summary, the present results indicate that the novel pro-inflammatory molecules S100A8/A9 are very early and sensitive markers of the exercise-induced inflammatory response. Further investigations are necessary to evaluate the applicability of S100A8/A9 for monitoring the training process and to elucidate the dependence on training status.

Fatty acids in serum cholesteryl esters in relation to asthma and lung function in children.

BACKGROUND: Dietary fatty acid intake has been proposed to contribute to asthma development with n-6 polyunsaturated fatty acids (PUFA) having a detrimental and n-3 PUFA a protective effect. OBJECTIVE: The aim of our analysis was to explore the relationship between fatty acid composition of serum cholesteryl esters as marker of dietary intake and prevalence of asthma, impaired lung function and bronchial hyper-responsiveness in children. METHODS: The study population consisted of 242 girls and 284 boys aged 8-11 years, living in Munich, Germany. Data were collected by parental questionnaire, lung function measurement and skin prick test according to the International Study of Asthma and Allergies in Childhood phase II protocol. Confounder-adjusted odds ratios (OR) with 95% confidence intervals (CI) were calculated for the association between quartiles of fatty acid concentration and health outcomes with the first quartile as reference. RESULTS: n-3 PUFA: levels of eicosapentaenoic acid were not related to asthma and impaired lung function. Linolenic acid levels were positively associated with current asthma (OR for fourth quartile 3.35, 95% CI 1.29-8.66). Forced expiratory volume in 1 s (FEV(1)) values decreased with increasing levels of linolenic acid (p for trend=0.057). n-6 PUFA: there was a strong positive association between arachidonic acid levels and current asthma (OR(4th quartile) 4.54, 1.77-11.62) and a negative association with FEV(1) (P=0.036). In contrast, linoleic acid was negatively related to current asthma (OR(4th quartile) 0.34, 0.14-0.87) and FEV(1) values increased with increasing levels of linoleic acid (P=0.022). The ratio of measured n-6 to n-3 PUFA as well as levels of palmitic and oleic acid were not consistently related to asthma or lung function. CONCLUSION: Our data do not support the hypothesis of a protective role of n-3 PUFA. Elevated arachidonic acid levels in children with asthma may be because of a disturbed balance in the metabolism of n-6 PUFA or may be secondary to inflammation in these patients.

Determination of cyclosporine and its metabolites in blood via HPLC-MS and correlation to clinically important parameters.

The narrow therapeutic window of the immunosuppressive drug cyclosporine (CsA), the interindividual variability of its metabolism, and the immunosuppressive activity/toxicity of some metabolites require investigation to correlate the parent substance and its metabolites and observed clinical parameters. Improved knowledge about these correlations may improve postoperative treatment of transplant patients. To observe such correlation therapeutic drug monitoring was performed by high-performance liquid chromatography-mass spectrometry (HPLC-MS) on 202 blood samples of kidney transplant patients. As CsA and its metabolites are preferably bound to lipoproteins in vivo, sample preparation included protein precipitation, solid phase extraction, and separation on a reversed phase column. Mass-spectrometric detection by an electrospray ionization chamber made the detection and quantification of the sodium adducts of CsA and its metabolites AM1, AM1c, DihydroAM1, AM19, and AM4N possible. With the presented HPLC-MS method, rapid information was achieved about the specific metabolization in a patient. Statistical computations related CsA and its metabolite concentrations to clinically important blood parameters. Significant correlation to the blood level of bilirubin and liver enzymes confirmed the presumed hepatotoxic potential of CsA and some metabolites. Furthermore, a strong correlation of AM19 to CRP and IL6 was observerd. These parameters may influence the prognosis for atherosclerosis, inflammation, and chronic allograft nephropathy.

Receptor assay based on surface plasmon resonance for the assessment of the complex formation activity of cyclosporin A and its metabolites.

OBJECTIVE: A new automated receptor assay has been used to determine the complex formation activity of cyclosporin A (CsA) and its metabolites in whole blood. METHODS: CsA in vivo forms a complex with cyclophilin A and calcineurin leading to an inhibition of the calmodulin-dependent phosphatase activity of calcineurin. The equilibrium complex formation gives information about the potential immunosuppressive activity of CsA and its metabolites. To measure the amount of this complex the authors developed an automated receptor assay based on an optical biosensor (Biacore) with surface plasmon resonance (SPR) technology. RESULTS: In the range of 50-300 nM CsA, the intra-day coefficient of variation (CV) was 7.2%, and the inter-day CV was 10.1%. Measuring range of the assay was 10-500 nM with a detection limit of 5 nM and a processing time of 10 min. Recovery rate for sample pretreatment was 74 +/- 5%. 193 blood specimens from heart transplant recipients were analyzed with 3 different methods. The results determined with the receptor assay were correlated with those obtained by fluorescence polarization immunoassay (FPIA; r = 0.599) and high-performance liquid chromatography (HPLC; r = 0.615). CONCLUSION: The receptor assay determines the complex formation activity of CsA and its metabolites with high sensitivity and precision.

Effects of dietary fatty acids on the composition and oxidizability of low-density lipoprotein.

OBJECTIVE: The objective of this study was to compare the effects of dietary monounsaturated fatty acids (MUFA), n-6 and n-3 polyunsaturated fatty acids (PUFA) on LDL composition and oxidizability. DESIGN, SETTING AND SUBJECTS: Sixty-nine healthy young volunteers, students at a nearby college, were included. Six subjects withdrew because of intercurrent illness and five withdrew because they were unable to comply with the dietary regimen. INTERVENTIONS: The participants received a 2-week wash-in diet rich in saturated fatty acids (SFA) followed by diets rich in refined olive oil, rapeseed oil or sunflower oil for 4 weeks. Intakes of vitamin E and other antioxidants did not differ significantly between the diets. RESULTS: At the end of the study, LDL oxidizability was lowest in the olive oil group (lag time: 72.6 min), intermediate in the rapeseed oil group (68.2 min) and highest in the sunflower oil group (60.4 min, P<0.05 for comparison of all three groups). Despite wide variations in SFA intake, the SFA content of LDL was not statistically different between the four diets (25.8-28.5% of LDL fatty acids). By contrast, the PUFA (43.5%-60.5% of LDL fatty acids) and MUFA content of LDL (13.7-29.1% of LDL fatty acids) showed a wider variability dependent on diet. CONCLUSIONS: Enrichment of LDL with MUFA reduces LDL susceptibility to oxidation. As seen on the rapeseed oil diet this effect is independent of a displacement of higher unsaturated fatty acids from LDL. Evidence from this diet also suggests that highly unsaturated n-3 fatty acids in moderate amounts do not increase LDL oxidizability when provided in the context of a diet rich in MUFA.

Associations between homocysteine and coagulation factors--a cross-sectional study in two populations of central Europe.

Plasma homocysteine has been associated with vascular disease and mortality. Experimental studies and studies on patients with vascular disease have indicated a thrombogenic potential of raised homocysteine levels. Studies on community samples are rare. We investigated the associations between homocysteine levels and selected coagulation factors in population-based random samples of 187 men from Pardubice (Czech Republic) and 147 men from Augsburg (Germany), aged 45 to 64 years. Czech men had higher mean levels of plasma homocysteine (10.3 vs. 8.9 micromol/l, P<.001) and of fibrinogen, von Willebrand factor (vWF), prothrombin fragment 1+2 (F 1+2) and D-Dimer (each P<.05). Plasma homocysteine was positively correlated with fibrinogen (r=.34) and vWF (r=.23, each P<.001) only in Czechs, and with D-Dimer in both Czechs and Germans (r=.26 and.21, respectively). Formal testing for interaction regarding the intercountry differences in the relationship with homocysteine revealed significance only for fibrinogen (P<.01). In multivariate analyses, the association of homocysteine with D-Dimer remained statistically significant after adjustment for indicators of chronic inflammation and fibrinogen. No significant correlation was found with Factor VII (F VII) activity or F 1+2. Homocysteine levels were also unrelated to traditional risk factors. In conclusion, in these cross-sectional studies we found moderate to strong associations between homocysteine and components of the endogenous hemostatic and fibrinolytic systems. The associations were slightly different between Czech and German men. These findings may help to better understand the role of homocysteine in atherothrombotic diseases.

Accumulation of cardiolipin and lysocardiolipin in fibroblasts from Tangier disease subjects.

Tangier disease (TD) is an inherited disorder of lipid metabolism characterized by very low high density lipoprotein (HDL) plasma levels, cellular cholesteryl ester accumulation and reduced cholesterol excretion in response to HDL apolipoproteins. Molecular defects in the ATP binding cassette transporter 1 (ABCA1) have recently been identified as the cause of TD. ABCA1 plays a key role in the translocation of cholesterol across the plasma membrane, and defective ABCA1 causes cholesterol storage in TD cells. Not only cholesterol efflux, but also phospholipid efflux was shown to be impaired in TD cells. By use of thin layer chromatography, high performance liquid chromatography and time-of-flight secondary ion mass spectrometry, we characterized the cellular phospholipid content in fibroblasts from three homozygous TD patients. The cellular content of the major phospholipids was not found to be significantly altered in TD fibroblasts. However, the two phospholipids cardiolipin and lysocardiolipin, which make up minute amounts in normal cells, were at least 3-5-fold enriched in fibroblasts from TD subjects. A structurally closely related phospholipid (lysobisphosphatidic acid) has recently been shown to be enriched in Niemann-Pick type C, another lipid storage disorder. Altogether these data may indicate that the role of these phospholipids is a regulatory one rather than that of a bulk mediator of cholesterol solubilization in sterol trafficking and efflux.

Suppression of endothelial cell apoptosis by high density lipoproteins (HDL) and HDL-associated lysosphingolipids.

Apoptotic cell death following injury of vascular endothelium is assumed to play an important role in the pathogenesis of atherosclerosis. In this report, we demonstrate that high density lipoproteins (HDL), a major anti-atherogenic lipoprotein fraction, protect endothelial cells against growth factor deprivation-induced apoptosis. HDL blocked the mitochondrial pathway of apoptosis by inhibiting dissipation of mitochondrial potential (Deltapsi(m)), generation of reactive oxygen species, and release of cytochrome c into the cytoplasm. As a consequence, HDL prevented activation of caspases 9 and 3 and apoptotic alterations of the plasma membrane such as increase of permeability and translocation of phosphatidylserine. Treatment of endothelial cells with HDL induced activation of the protein kinase Akt, an ubiquitous transducer of anti-apoptotic signals, and led to phosphorylation of BAD, a major Akt substrate. Suppression of Akt activity both by wortmannin and LY-294002 or by a dominant negative Akt mutant abolished the anti-apoptotic effect of HDL. Two bioactive lysosphingolipids present in HDL particles, sphingosylphosphorylcholine and lysosulfatide, fully mimicked the survival effect of HDL by blocking the mitochondrial pathway of apoptosis and potently activating Akt. In conclusion, the present study identifies HDL as a carrier of endogenous endothelial survival factors and suggests that inhibition of endothelial apoptosis by HDL-associated lysosphingolipids may represent an important and novel aspect of the anti-atherogenic activity of HDL.

High density lipoproteins induce cell cycle entry in vascular smooth muscle cells via mitogen activated protein kinase-dependent pathway.

In this study we found that HDL acts as a potent and specific mitogen in vascular smooth muscle cells (VSMC) by stimulating entry into S-phase and DNA synthesis in a time- and concentration-dependent manner, induction of cyclins D1, E, and A, as well as activation of cyclin D-dependent kinases as inferred from phosphorylation of the retinoblastoma protein (pRb). Moreover, HDL induced activation of the mitogen-activated protein kinase pathway including Raf-, MEK-1, and ERK1/2, as well as the expression of proto-oncogen c-fos, which is controlled by ERK1/2. PD98059, an inhibitor of MEK-1 blocked the mitogenic activity of HDL and cyclin D1 expression. HDL-induced VSMC proliferation, cell cycle progression, cyclin D1 expression, and activation of the Raf-1/MEK-1/ERK1/2 cascade were blocked by preincubation of cells with pertussis toxin indicating involvement of trimeric G-protein. By contrast, none of these responses was inhibited by the protein kinase C inhibitor, GF109203X. The mitogenic effects of native HDL were not mimicked by apo A-I, reconstituted HDL containing apo A-I, or cholesterol-containing liposomes. In conclusion, HDL possesses an intrinsic property to induce G-protein- and MAP-kinase-dependent proliferation and cell cycle progression in VSMC. The strong and specific mitogenic effect of HDL should be taken into account, when therapeutic strategies to elevate the plasma level of these lipoproteins are developed.

Expression of the ATP-binding cassette transporter gene ABCG1 (ABC8) in Tangier disease.

Several members of the ATP-binding cassette (ABC) transporter family are involved in cholesterol efflux from cells. A defect in one member, ABCA1, results in Tangier disease, a condition characterized by cholesterol accumulation in macrophages and virtual absence of mature circulating high-density lipoproteins. Expression of a second member, ABCG1, is increased by cholesterol-loading in human macrophages. We now show that ABCG1, which we identified by differential display RT-PCR in foamy macrophages, is overexpressed in macrophages from patients with Tangier disease compared to control macrophages. On examination by confocal laser scanning microscopy, ABCG1 was present in perinuclear structures within the cell. In addition, a combination of in situ hybridization and indirect immunofluorescence microscopy revealed that ABCG1 is expressed in foamy macrophages within the atherosclerotic plaque. These data indicate that not only ABCA1 but also ABCG1 may play a role in the cholesterol metabolism of macrophages in vitro and in the atherosclerotic plaque.

Multimodal perioperative management--combining thoracic epidural analgesia, forced mobilization, and oral nutrition--reduces hormonal and metabolic stress and improves convalescence after major urologic surgery.

We sought in this prospective study to use a multimodal approach to reduce stress and improve recovery in patients undergoing major surgery. During an initial study period, 30 patients were randomly allocated to receive general anesthesia (GA; Group 1) or a combination of GA and intraoperative thoracic epidural analgesia (TEA; Group 2) when undergoing radical cystectomy. Parenteral nutrition was provided for 5 days after surgery. During the second period, 15 patients were treated with a multimodal approach (Group 3) consisting of intraoperative GA and TEA, postoperative patient-controlled TEA, early oral nutrition, and enforced mobilization. Data for plasma and urine catecholamines, plasma cortisol, the nitrogen balance, the postoperative inflammatory nutrition index, pain relief, fatigue, sleep, overnight recovery, recovery of bowel function, and mobilization were recorded up to the fifth postoperative day. Plasma concentrations of catecholamines and cortisol were comparable in all patients, but those in Group 3 had lower levels of urinary catecholamine excretion. Protein intake was more effective with parenteral nutrition. Nitrogen balances were less negative, and the postoperative inflammatory nutrition index score increased significantly in the traditional groups but not in Group 3. Multimodally treated patients reported less fatigue and better overnight recovery. Along with improved pain relief, recovery of bowel function, and ambulation, there were no differences in the postoperative complication rates among the three groups. The multimodal approach reduced stress and improved metabolism and recovery after radical cystectomy.