Early pharmacotherapy with fluoxetine rescues dendritic pathology in the Ts65Dn mouse model of down syndrome.

Down syndrome DS is a genetic pathology characterized by brain hypotrophy and severe cognitive impairment. Although defective neurogenesis is an important determinant of mental disability, a severe dendritic pathology appears to be an equally important factor. A previous study showed that fluoxetine, a selective serotonin reuptake inhibitor, fully restores neurogenesis in the Ts65Dn mouse model of DS. The goal of the current study was to establish whether fluoxetine also restores dendritic development. In mice aged 45 days, treated with fluoxetine in the postnatal period P3-P15, we examined the dendritic arbor of the granule cells of the dentate gyrus (DG). The granule cells of trisomic mice had a severely hypotrophic dendritic arbor, fewer spines and a reduced innervation than euploid mice. Treatment with fluoxetine fully restored all these defects. In Ts65Dn mice, we found reduced levels of serotonin that were restored by treatment. Results show that a pharmacotherapy with fluoxetine is able to rescue not only the number of granule neurons but also their "quality" in terms of correct maturation and connectivity. These findings strongly suggest that fluoxetine may be a drug of choice for the improvement of the major defects in the DS brain and, possibly, of mental retardation.

Statin therapy decreases serum levels of high-sensitivity C-reactive protein and tumor necrosis factor-α in HIV-infected patients treated with ritonavir-boosted protease inhibitors.

BACKGROUND: Statins are lipid-lowering drugs that exhibit anti-inflammatory and immune-modulatory properties, leading to a reduction of serum levels of C-reactive protein (CRP) in the general population. OBJECTIVE: Because very limited data are available today, our objective was to assess the lipid-lowering effects of statins and their capacity to decrease selected soluble markers of inflammation in HIV-infected patients. METHODS: Retrospective cohort study of HIV-infected adult patients with hypercholesterolemia who were receiving a stable antiretroviral regimen including a ritonavir-boosted protease inhibitor and who started a lipid-lowering therapy with rosuvastatin (10 mg daily), atorvastatin (10 mg daily), or pravastatin (40 mg daily) and were followed-up for at least 12 months. One hundred and fifty-one patients were enrolled in the study: 51 in the rosuvastatin group, 47 in the atorvastatin group, and 53 in the pravastatin group. The primary observation was change in plasma lipid levels and serum markers of inflammation (high-sensitivity C-reactive protein [hsCRP], interleukin-6 [IL-6], and tumor necrosis factor-α [TNF- α]), while secondary observations include immunovirological parameters and safety profile of statins. RESULTS: One year after starting the statin therapy, patients treated with rosuvastatin had significantly greater decreases in total cholesterol and LDL cholesterol than subjects on atorvastatin or pravastatin. All statins led to a similar, significant reduction in serum levels of hsCRP and TNF-α, without correlation between biomarkers and lipid values, and toxicity rates were similar for all 3 statins. CONCLUSION: Our findings suggest that rosuvastatin has a significantly greater lipid-lowering effect than atorvastatin or pravastatin, but all 3 statins exert a similar effect in lowering markers of inflammation as hsCRP and TNF-α.

Lopinavir/ritonavir trough concentrations with the tablet formulation in HIV-1-infected women during the third trimester of pregnancy.

OBJECTIVES: An observational, open-label study was performed to assess changes of lopinavir/ritonavir plasma concentrations during pregnancy. METHODS: Adult HIV-1-infected women during the third trimester of pregnancy and on stable antiretroviral treatment including zidovudine/lamivudine plus lopinavir/ritonavir tablets (400/100 mg twice daily) were asked to participate. This group was compared with a group of non-pregnant HIV-1-infected women receiving the same antiretroviral regimen. The trough plasma concentration (C(trough)) of lopinavir and ritonavir was assessed at steady-state by a validated high-performance liquid chromatography (HPLC)-tandem mass spectrometry method. RESULTS: A total of 41 HIV-positive female patients were enrolled in the study, with a median age of 28 y (range 20-37 y). These patients were stratified into 2 groups: 21 women in the third trimester of pregnancy (group A) and 20 non-pregnant women (group B). The geometric mean (95% confidence interval (CI)) plasma C(trough) of lopinavir was 4205 (2418-6896) ng/ml in group A and 5098 (3187-8084) ng/ml in group B. The reduction in lopinavir plasma levels observed in group A was not significant (geometric mean ratio 0.87, 95% CI 0.62-1.32; p = 0.411). No correlation was found between lopinavir plasma levels and adverse events (such as diarrhoea and hyperlipidaemia) or immunological parameters of HIV disease, and no changes in plasma HIV viral load were reported. CONCLUSION: In this study, a slight but not significant decrease in the plasma lopinavir C(trough) was found during the third trimester of pregnancy, suggesting that standard dosing of the tablet formulation is also appropriate during the later stages of pregnancy.

Estimation of reference intervals of five endocannabinoids and endocannabinoid related compounds in human plasma by two dimensional-LC/MS/MS.

The elucidation of the role of endocannabinoids in physiological and pathological conditions and the transferability of the importance of these mediators from basic evidence into clinical practice is still hampered by the indefiniteness of their circulating reference intervals. In this work, we developed and validated a two-dimensional LC/MS/MS method for the simultaneous measurement of plasma endocannabinoids and related compounds such as arachidonoyl-ethanolamide, palmitoyl-ethanolamide, and oleoyl-ethanolamide, belonging to the N-acyl-ethanolamide (NAE) family, and 2-arachidonoyl-glycerol and its inactive isomer 1-arachidonoyl-glycerol from the monoacyl-glycerol (MAG) family. We found that several pitfalls in the endocannabinoid measurement may occur, from blood withdrawal to plasma processing. Plasma extraction with toluene followed by on-line purification was chosen, allowing high-throughput and reliability. We estimated gender-specific reference intervals on 121 healthy normal weight subjects fulfilling rigorous anthropometric and hematic criteria. We observed no gender differences for NAEs, whereas significantly higher MAG levels were found in males compared with females. MAGs also significantly correlated with triglycerides. NAEs increased with age in females, and arachidonoyl-ethanolamide correlated with adiposity and metabolic parameters in females. This work paves the way to the establishment of definitive reference intervals for circulating endocannabinoids to help physicians move from the speculative research field into the clinical field.

Assessing the impact of hepatitis C virus coinfection on lopinavir/ritonavir trough concentrations in HIV-infected patients.

PURPOSE: Chronic hepatitis C is an emerging issue in the management of human immunodeficiency virus (HIV) disease because both diseases have the same route of transmission, leading to a very high prevalence of hepatitis C virus (HCV)-coinfection in the HIV-positive patient population. Lopinavir is extensively metabolized by the hepatic cytochrome P450 3A4, and the pharmacokinetics of this protease inhibitor (PI) could be influenced by liver impairment. However, data currently available on the impact of HCV-coinfection on lopinavir plasma concentrations are both limited and conflicting. METHODS: This was an observational, open-label study in which adult HIV-infected outpatients on stable antiretroviral treatment that included two nucleoside reverse transcriptase inhibitors (NRTIs) plus lopinavir/ritonavir for at least 4 weeks were asked to participate. The trough plasma concentration (C (trough)) of lopinavir and ritonavir was assessed at steady state by a validated high-performance liquid chromatography-tandem mass spectrometry method. RESULTS: A total of 65 HIV-positive patients were enrolled in the study. These patients were stratified into two groups based on the absence/presence of HCV-coinfection: 45 were monoinfected (HIV+/HCV-) and 20 were coinfected (HIV+/HCV+). The lopinavir C (trough) in plasma was comparable between HIV+/HCV+ and HIV+/HCV- patients, without any statistically significant difference (geometric mean ratio 0.89, 95% confidence interval 0.61-1.42; p = 0.581). The mean ritonavir C (trough) was also comparable in the two groups. Almost all samples were found to be within the therapeutic plasma level range (97% in HIV+/HCV- group and 100% in HIV+/HCV+ group). No correlation was found between lopinavir plasma levels and adverse events (such as diarrhoea and hypertriglyceridaemia) or immune-virological parameters of HIV disease. CONCLUSIONS: Among the HIV-positive patients participating in this study, the pharmacokinetics of lopinavir/ritonavir did not significantly change in those HIV-positive patients coinfected with HCV and in the absence of liver cirrhosis.

Serum steroid profiling by isotopic dilution-liquid chromatography-mass spectrometry: comparison with current immunoassays and reference intervals in healthy adults.

BACKGROUND: The simultaneous, rapid and reliable measurement of a wide steroid panel is a powerful tool to unravel physiological and pathological hormone status. Clinical laboratories are currently dominated by high-throughput immunoassays, but these methods lack specificity due to cross-reactivity and matrix interferences. We developed and validated an isotopic dilution-liquid chromatography-tandem mass spectrometry (ID-LC-MS/MS) method for the simultaneous measurement of cortisol, corticosterone, 11deoxycortisol, androstenedione, deoxycorticosterone (DOC), testosterone, 17OHprogesterone, dehydroepiandrosterone (DHEA) and progesterone in serum, and compared it to routine immunoassays employed in our laboratory. We also established adult reference intervals in 416 healthy subjects. METHODS: 0.9 ml of serum were spiked with labelled internal standards (IS) and extracted on C18 cartridges. Eluate was injected into a two-dimensional LC-system, purified in a perfusion column and separated on a C8 column during a 21 min gradient run. Analytes were revealed by atmospheric pressure chemical ionization (APCI) followed by multiple reaction monitoring (MRM) analysis. RESULTS: Of the four immunoassays compared with the ID-LC-MS/MS method, only the results of ElecsysE170 for cortisol, testosterone in males and progesterone>1 ng/ml were in agreement with ID-LC-MS/MS. ElecsysE170 for testosterone in females and progesterone<1 ng/ml, Immulite2000 for androstenedione, DSL-9000 for DHEA and 17OHP Bridge for 17OHprogesterone, respectively, showed poor agreement. Reference intervals and steroid age and fertility related fluctuations were established. CONCLUSION: Our ID-LC-MS/MS method proved to be reliable and sensitive in revealing steroid circulating concentrations in adults and in highlighting the limits of routine immunoassays at low concentrations.

Early pharmacotherapy restores neurogenesis and cognitive performance in the Ts65Dn mouse model for Down syndrome.

Down syndrome (DS) is a genetic pathology characterized by intellectual disability and brain hypotrophy. Widespread neurogenesis impairment characterizes the fetal and neonatal DS brain, strongly suggesting that this defect may be a major determinant of mental retardation. Our goal was to establish, in a mouse model for DS, whether early pharmacotherapy improves neurogenesis and cognitive behavior. Neonate Ts65Dn mice were treated from postnatal day (P) 3 to P15 with fluoxetine, an antidepressant that inhibits serotonin (5-HT) reuptake and increases proliferation in the adult Ts65Dn mouse (Clark et al., 2006). On P15, they received a BrdU injection and were killed after either 2 h or 1 month. Results showed that P15 Ts65Dn mice had notably defective proliferation in the hippocampal dentate gyrus, subventricular zone, striatum, and neocortex and that proliferation was completely rescued by fluoxetine. In the hippocampus of untreated P15 Ts65Dn mice, we found normal 5-HT levels but a lower expression of 5-HT1A receptors and brain-derived neurotrophic factor (BDNF). In Ts65Dn mice, fluoxetine treatment restored the expression of 5-HT1A receptors and BDNF. One month after cessation of treatment, there were more surviving cells in the dentate gyrus of Ts65Dn mice, more cells with a neuronal phenotype, more proliferating precursors, and more granule cells. These animals were tested for contextual fear conditioning, a hippocampus-dependent memory task, and exhibited a complete recovery of memory performance. Results show that early pharmacotherapy with a drug usable by humans can correct neurogenesis and behavioral impairment in a model for DS.

The spectrum of plant and animal sterols in different oil-derived intravenous emulsions.

Intravenous lipid constituents have different effects on various biological processes. Some of these effects are protective, while others are potentially adverse. Phytosterols, in particular, seem to be implicated with parenteral nutrition-associated cholestasis. The aim of this study is to determine the amount of plant and animal sterols present in lipid formulations derived from different oil sources. To this end, animal (cholesterol) and plant (beta-sitosterol, campesterol, and stigmasterol) sterols in seven different commercially available intravenous lipid emulsions (ILEs) were quantified by capillary gas chromatography after performing a lipid extraction procedure. The two major constituents of the lipid emulsions were cholesterol (range 14-57% of total lipids) and beta-sitosterol (range 24-55%), followed by campesterol (range 8-18%) and stigmasterol (range 5-16%). The fish oil-derived formulation was an exception, as it contained only cholesterol. The mean values of the different sterols were statistically different across ILEs (P = 0.0000). A large percentage of pairwise comparisons were also statistically significant (P = 0.000), most notably for cholesterol and stigmasterol (14 out of 21 for both), followed by campesterol (12 out 21) and beta-sitosterol (11 out 21). In conclusion, most ILEs combined significant amounts of phytosterols and cholesterol. However, their phytosterols:cholesterol ratios were reversed compared to the normal human diet.

Vitamin A and vitamin E isoforms stability and peroxidation potential of all-in-one admixtures for parenteral nutrition.

BACKGROUND: In all-in-one admixtures (AIOs), vitamins can be degraded and lipid can be peroxidized by light exposure, oxygen action, and multiple chemical interactions. AIM: We investigated the impact of three commercial lipid emulsions and two multivitamin preparations on vitamin A and vitamin E chemical stability and lipid peroxidation potential of AIOs. METHODS: A soybean oil (Soy), soybean/medium-chain triacylglycerol oil (MCT), and olive/soybean oil (Olive)-based emulsion (all 20%), and a lyophilized (Lyo) and emulsified (Emu) multivitamin compounds, were tested. Two AIOs for each lipid emulsion were prepared, the former with Lyo and the latter with Emu. The concentrations of retinol palmitate, alpha-gamma-delta-tocopherol, and malondialdehyde were analyzed in AIOs, immediately (T0) and 24 hours (T24) after compounding. RESULTS: Retinol palmitate, and alpha- and gamma-tocopherol were more stable in MCT-AIOs than in both Soy-AIOs and Olive-AIOs (p < 0.013; p < 0.001 respectively). Furthermore alpha-tocopherol was more stable in Lyo-AIOs than in Emu-AIOs (p < 0.004). Malondialdehyde (MDA) increased differently among the admixtures; however the concentrations were similar in all AIOs at T24. CONCLUSIONS: The differences in retinol palmitate stability were due both to lipid emulsions per se and to interaction between lipid emulsions and multivitamin preparations. The alpha-gamma-tocopherol stability depended on both lipid emulsions and multivitamin preparations. In tested AIOs there was a different degradation rate of fat-soluble vitamins to keep the same lipid peroxidation level, since MDA concentrations at T24 were similar among AIOs.

5-methyltetrahydrofolate administration is associated with prolonged survival and reduced inflammation in ESRD patients.

BACKGROUND: Hemodialysis (HD) patients have a greatly increased risk of cardiovascular morbidity and mortality. For this reason, attempts are often made to normalize hyperhomocysteinemia. This randomized prospective study sought to determine which risk factors are predictors of mortality and whether high doses of folates or 5-methyltetrahydrofolate (5-MTHF) could improve hyperhomocysteinemia and survival in HD patients. METHODS: 341 patients were divided into two groups: group A was treated with 50 mg i.v. 5-MTHF, and group B was treated with 5 mg/day oral folic acid. Both groups received i.v. vitamin B(6) and B(12). By dividing patients into C-reactive protein (CRP) quartiles, group A had the highest survival for CRP <12 mg/l, whereas no survival difference was found for group B. CRP was the only predictive risk factor for death (RR 1.17, range 1.04-1.30, p = 0.02). Dialysis age, hyperhomocysteinemia, methylenetetrahydrofolate reductase polymorphism, albumin, lipoprotein (a) and folate did not influence mortality risk. Survival in group A was higher than that in group B, namely 36.2 +/- 20.9 vs. 26.1 +/- 22.2 months (p = 0.003). RESULTS: Our results suggest that CRP, but not hyperhomocysteinemia, is the main risk factor for mortality in HD patients receiving vitamin supplements. Intravenous 5-MTHF seems to improve survival in HD patients independent from homocysteine lowering.

Influence of age, sex and vitamin status on fasting and post-methionine load plasma homocysteine levels.

BACKGROUND AND OBJECTIVES: To investigate the effects of age, sex and vitamin status on total plasma homocysteine (tHCy), both after fasting (FtHCy) and two hours post-methionine load (PML-tHCy). The secondary aim was to determine the reference values for FtHCy and PML-tHCy. DESIGN AND METHODS: A cohort of apparently healthy volunteers underwent blood sampling for FtHCy, PML-tHCy, creatinine, serum folate, vitamin B12 and vitamin B6 (pyridoxal-5-phosphate, PLP). RESULTS: In 147 subjects (M/F= 82/65, age range: 14-94 years), FtHCy was significantly higher in men than in women. In men, age and folate levels explained 20.5% and 19.0% of FtHCy variance, respectively. In women, age and vitamin B12 accounted for 22.6% and 17.8% of FtHCy variance, respectively. PML-tHCy was similar in men and women. PML-tHCy was negatively correlated with folate in both sexes, and with vitamin B12 and age in women only. Folate accounted for 20% of the variance of PML-tHCy in men, while in women vitamin B12 and PLP explained 40% and 20% of variance of PML-tHCy, respectively. The reference values of FtHCy and PML-tHCy were: 19.63 and 40.18 mol/L, respectively, for men under 45 years, 14.26 and 28.31 mol/L, respectively, for women under 45 years, 28.38 and 36.48 mol/L for men above 45 years, and 22.49 and 44.06 mol/L for women above 45 years. INTERPRETATION AND CONCLUSIONS: Age, gender and vitamin status influence both FtHCy and PML-tHCy in normal subjects. Reference values should be calculated according to age and sex.