Protective effect of intravitreal administration of tresperimus, an immunosuppressive drug, on experimental autoimmune uveoretinitis.

PURPOSE: To test the efficiency of locally administrated tresperimus in experimental autoimmune uveoretinitis (EAU). METHODS: EAU was induced in Lewis rats by S-antigen (S-Ag) immunization. Three intravitreal injections of tresperimus (prevention or prevention/treatment protocols) were performed at different time points after immunization. The pharmacokinetics of tresperimus was evaluated in the ocular tissues and plasma. The in vitro effect of tresperimus was evaluated on macrophages. EAU was graded clinically and histologically. Blood ocular barrier permeability was evaluated by protein concentration in ocular fluids. Immune response to S-Ag was examined by delayed type hypersensitivity, the expression of inflammatory cytokines in lymph nodes, ocular fluids and serum by multiplex ELISA, and in ocular cells by RT-PCR. RESULTS: In vitro, tresperimus significantly reduced the production of inflammatory cytokines by lipopolysaccharide-stimulated macrophages. In vivo, in the treatment protocol, efficient tresperimus levels were measured in the eye but not in the plasma up to 8 days after the last injection. Tresperimus efficiently reduced inflammation, retinal damage, and blood ocular barrier permeability breakdown. It inhibited nitric oxide synthase-2 and nuclear factor κBp65 expression in ocular macrophages. IL-2 and IL-17 were decreased in ocular media, while IL-18 was increased. By contrast, IL-2 and IL-17 levels were not modified in inguinal lymph nodes draining the immunization site. Moreover, cytokine levels in serum and delayed type hypersensitivity to S-Ag were not different in control and treated rats. In the prevention/treatment protocol, ocular immunosuppressive effects were also observed. CONCLUSIONS: Locally administered tresperimus appears to be a potential immunosuppressive agent in the management of intraocular inflammation.

Tenofovir use is associated with a reduction in calculated glomerular filtration rates in the Swiss HIV Cohort Study.

BACKGROUND: A growing number of case reports have described tenofovir (TDF)-related proximal renal tubulopathy and impaired calculated glomerular filtration rates (cGFR). We assessed TDF-associated changes in cGFR in a large observational HIV cohort. METHODS: We compared treatment-naive patients or patients with treatment interruptions > or = 12 months starting either a TDF-based combination antiretroviral therapy (cART) (n = 363) or a TDF-sparing regime (n = 715). The predefined primary endpoint was the time to a 10 ml/min reduction in cGFR, based on the Cockcroft-Gault equation, confirmed by a follow-up measurement at least 1 month later. In sensitivity analyses, secondary endpoints including calculations based on the modified diet in renal disease (MDRD) formula were considered. Endpoints were modelled using pre-specified covariates in a multiple Cox proportional hazards model. RESULTS: Two-year event-free probabilities were 0.65 (95% confidence interval [CI] 0.58-0.72) and 0.80 (95% CI 0.76-0.83) for patients starting TDF-containing or TDF-sparing cART, respectively. In the multiple Cox model, diabetes mellitus (hazard ratio [HR] = 2.34 [95% CI 1.24-4.42]), higher baseline cGFR (HR = 1.03 [95% CI 1.02-1.04] by 10 ml/min), TDF use (HR = 1.84 [95% CI 1.35-2.51]) and boosted protease inhibitor use (HR = 1.71 [95% CI 1.30-2.24]) significantly increased the risk for reaching the primary endpoint. Sensitivity analyses showed high consistency. CONCLUSION: There is consistent evidence for a significant reduction in cGFR associated with TDF use in HIV-infected patients. Our findings call for a strict monitoring of renal function in long-term TDF users with tests that distinguish between glomerular dysfunction and proximal renal tubulopathy, a known adverse effect of TDF.

Effects of the α₂-adrenergic agonist clonidine on the pharmacodynamics and pharmacokinetics of 3,4-methylenedioxymethamphetamine in healthy volunteers.

The mechanism of action of 3,4-methylenedioxymethamphetamine (MDMA; ecstasy) involves the carrier-mediated and potentially vesicular release of monoamines. We assessed the effects of the sympatholytic α₂-adrenergic receptor agonist clonidine (150 μg p.o.), which inhibits the neuronal vesicular release of norepinephrine, on the cardiovascular and psychotropic response to MDMA (125 mg p.o.) in 16 healthy subjects. The study used a randomized, double-blind, placebo-controlled crossover design with four experimental sessions. The administration of clonidine 1 h before MDMA reduced the MDMA-induced increases in plasma norepinephrine concentrations and blood pressure but only to the extent that clonidine lowered norepinephrine levels and blood pressure compared with placebo. Thus, no interaction was found between the cardiovascular effects of the two drugs. Clonidine did not affect the psychotropic effects or pharmacokinetics of MDMA. The lack of an interaction of the effects of clonidine and MDMA indicates that vesicular release of norepinephrine, which is inhibited by clonidine, does not critically contribute to the effects of MDMA in humans. Although clonidine may be used in the treatment of stimulant-induced hypertensive reactions, the present findings do not support a role for α₂-adrenergic receptor agonists in the prevention of psychostimulant dependence.

Blunting the response to endotoxin in healthy subjects: effects of various doses of intravenous fish oil.

To test the dose response effect of infused fish oil (FO) rich in n-3 PUFAs on the inflammatory response to endotoxin (LPS) and on membrane incorporation of fatty acids in healthy subjects. Prospective, sequential investigation comparing three different FO doses. Three groups of male subjects aged 26.8 +/- 3.2 years (BMI 22.5 +/- 2.1). One of three FO doses (Omegaven10%) as a slow infusion before LPS: 0.5 g/kg 1 day before LPS, 0.2 g/kg 1 day before, or 0.2 g/kg 2 h before. Temperature, hemodynamic variables, indirect calorimetry and blood samples (TNF-alpha, stress hormones) were collected. After LPS temperature, ACTH and TNF-alpha concentrations increased in the three groups: the responses were significantly blunted (p < 0.0001) compared with the control group of the Pluess et al. trial. Cortisol was unchanged. Lowest plasma ACTH, TNF-alpha and temperature AUC values were observed after a single 0.2 g/kg dose of FO. EPA incorporation into platelet membranes was dose-dependent. Having previously shown that the response to LPS was reproducible, this study shows that three FO doses blunted it to various degrees. The 0.2 g/kg perfusion immediately before LPS was the most efficient in blunting the responses, suggesting LPS capture in addition to the systemic and membrane effects.

A modular approach for integrative analysis of large-scale gene-expression and drug-response data

High-throughput technologies are now used to generate more than one type of data from the same biological samples. To properly integrate such data, we propose using co-modules, which describe coherent patterns across paired data sets, and conceive several modular methods for their identification. We first test these methods using in silico data, demonstrating that the integrative scheme of our Ping-Pong Algorithm uncovers drug-gene associations more accurately when considering noisy or complex data. Second, we provide an extensive comparative study using the gene-expression and drug-response data from the NCI-60 cell lines. Using information from the DrugBank and the Connectivity Map databases we show that the Ping-Pong Algorithm predicts drug-gene associations significantly better than other methods. Co-modules provide insights into possible mechanisms of action for a wide range of drugs and suggest new targets for therapy

Substitution of (R,S)-methadone by (R)-methadone: Impact on QTc interval.

Methadone is administered as a chiral mixture of (R,S)-methadone. The opioid effect is mainly mediated by (R)-methadone, whereas (S)-methadone blocks the human ether-à-go-go-related gene (hERG) voltage-gated potassium channel more potently, which can cause drug-induced long QT syndrome, leading to potentially lethal ventricular tachyarrhythmias. To investigate whether substitution of (R,S)-methadone by (R)-methadone could reduce the corrected QT (QTc) interval, (R,S)-methadone was replaced by (R)-methadone (half-dose) in 39 opioid-dependent patients receiving maintenance treatment for 14 days. (R)-methadone was then replaced by the initial dose of (R,S)-methadone for 14 days (n = 29). Trough (R)-methadone and (S)-methadone plasma levels and electrocardiogram measurements were taken. The Fridericia-corrected QT (QTcF) interval decreased when (R,S)-methadone was replaced by a half-dose of (R)-methadone; the median (interquartile range [IQR]) values were 423 (398-440) milliseconds (ms) and 412 (395-431) ms (P = .06) at days 0 and 14, respectively. Using a univariate mixed-effect linear model, the QTcF value decreased by a mean of -3.9 ms (95% confidence interval [CI], -7.7 to -0.2) per week (P = .04). The QTcF value increased when (R)-methadone was replaced by the initial dose of (R,S)-methadone for 14 days; median (IQR) values were 424 (398-436) ms and 424 (412-443) ms (P = .01) at days 14 and 28, respectively. The univariate model showed that the QTcF value increased by a mean of 4.7 ms (95% CI, 1.3-8.1) per week (P = .006). Substitution of (R,S)-methadone by (R)-methadone reduces the QTc interval value. A safer cardiac profile of (R)-methadone is in agreement with previous in vitro and pharmacogenetic studies. If the present results are confirmed by larger studies, (R)-methadone should be prescribed instead of (R,S)-methadone to reduce the risk of cardiac toxic effects and sudden death.

Metabolic effects of parenteral nutrition enriched with n-3 polyunsaturated fatty acids in critically ill patients

BACKGROUND & AIMS: n-3 fatty acids are expected to downregulate the inflammatory responses, and hence may decrease insulin resistance. On the other hand, n-3 fatty acid supplementation has been reported to increase glycemia in type 2 diabetes. We therefore assessed the effect of n-3 fatty acids delivered with parenteral nutrition on glucose metabolism in surgical intensive care patients. METHODS: Twenty-four surgical intensive care patients were randomized to receive parenteral nutrition providing 1.25 times their fasting energy expenditure, with 0.25 g of either an n-3 fatty acid enriched-or a soy bean-lipid emulsion. Energy metabolism, glucose production, gluconeogenesis and hepatic de novo lipogenesis were evaluated after 4 days. RESULTS: Total energy expenditure was significantly lower in patients receiving n-3 fatty acids (0.015+/-0.001 vs. 0.019+/-0.001 kcal/kg/min with soy bean lipids (P<0.05)). Glucose oxidation, lipid oxidation, glucose production, gluconeogenesis, hepatic de novo lipogenesis, plasma glucose, insulin and glucagon concentrations did not differ (all P>0.05) in the 2 groups. CONCLUSIONS: n-3 fatty acids were well tolerated in this group of severely ill patients. They decreased total energy expenditure without adverse metabolic effects.

L'angioplastie coronaire chez le patient âgé [Coronary angioplasty in elderly patients]

Percutaneous transluminal coronary angioplasty (PTCA) is a widely accepted treatment of symptomatic coronary heart disease providing prompt and prolonged clinical, improvement in most patients. We have examined the value of this therapy in a group of 91 patients in their eighth decade treated by 133 consecutive angioplasties. Most patients had refractory or instable angor in spite of optimal pharmacotherapy. Multivessel disease was present in 67% and maintained left-ventricular function in 92% of the patients. The angiographic success rate of PTCA was 84%; technical failures occurred in 12% and adverse events in 14%. Two patients died. The rate of symptomatic restenosis was 24%. Survival and patients free of myocardial events were at 89% and 60%, respectively, estimated by Kaplan-Meier analysis. PTCA is an efficient and acceptable treatment for the elderly patient with severe and drug-resistant angina. Two years after PTCA the majority of patients was asymptomatic.

Role of glutathione in macrophage activation: effect of cellular glutathione depletion on nitrite production and leishmanicidal activity.

We have examined the effects of two agents depleting the intracellular pool of glutathione (GSH) on macrophage activation induced by IFN-gamma + LPS, as measured by nitrite production and leishmanicidal activity. Diethylmaleate (DEM), which depletes intracellular GSH by conjugation via a reaction catalyzed by the GSH-S-transferase, strongly inhibited nitrite secretion and leishmanicidal activity when added before or at the time of addition of IFN-gamma + LPS; this inhibition was progressively lost when addition of DEM was delayed up to 10 hr. A close correlation was observed between levels of intracellular soluble GSH during activation and nitrite secretion. Inhibition was partially reversed by the addition of glutathione ethyl ester (GSH-Et). Buthionine sulfoximine (BSO), a specific inhibitor of gamma-glutamylcysteine synthetase, also inhibited macrophage activation, although to a lesser extent than DEM despite a more pronounced soluble GSH depletion. This inhibition was completely reversed by the addition of GSH-Et. DEM and BSO did not alter cell viability or PMA-triggered O2- production by activated macrophages, suggesting that the inhibitory effects observed on nitrite secretion and leishmanicidal activity were not related to a general impairment of macrophage function. DEM and BSO treatment reduced iNOS specific activity and iNOS protein in cytosolic extracts. DEM also decreased iNOS mRNA expression while BSO had no effect. Although commonly used as a GSH-depleting agent, DEM may have additional effects because it can also act as a sulhydryl reagent; BSO, on the other hand, which depletes GSH by enzymatic inhibition, has no effect on protein-bound GSH. Our results suggest that both soluble and protein-bound GSH may be important for the induction of NO synthase in IFN-gamma + LPS-activated macrophages.

Deficient T cell fate specification in mice with an induced inactivation of Notch1.

Notch proteins are cell surface receptors that mediate developmental cell specification events. To explore the function of murine Notch1, an essential portion of the gene was flanked with loxP sites and inactivation induced via interferon-regulated Cre recombinase. Mice with a neonatally induced loss of Notch1 function were transiently growth retarded and had a severe deficiency in thymocyte development. Competitive repopulation of lethally irradiated wild-type hosts with wild-type- and Notch1-deficient bone marrow revealed a cell autonomous blockage in T cell development at an early stage, before expression of T cell lineage markers. Notch1-deficient bone marrow did, however, contribute normally to all other hematopoietic lineages. These findings suggest that Notch1 plays an obligatory and selective role in T cell lineage induction.

Thermogenic effects of commercially available plant preparations aimed at treating human obesity.

Different commercially available plant preparations have been claimed to have anti-obesity action. We investigated the acute effects of oral administration of 12 of these preparations in non-obese women and men. No significant increase in energy expenditure (EE) has been noted after treatment with any of these preparations. No change in respiratory quotient (RQ) was shown, except after treatment with maté (Ilex paraguariensis) extract, where a drop in RQ was observed, indicating a rise in the proportion of fat oxidized. The results suggested the poor potential of these plant preparations in the treatment of obesity, except possibly for the maté extract. Further studies are required to explore the influence of higher dosages of these preparations as well as chronic administration in man.

Effects of corticosterone on innate and humoral immune functions and oxidative stress in barn owl nestlings.

The costs of coping with stressful situations are traded-off against other functions such as immune responses. This trade-off may explain why corticosterone secretion reduces immune reactions. Corticosterone differentially affects various immunity components. However, which component is suppressed varies between studies. It remains unclear whether the trade-off in energy, nutrition, autoimmunity or oxidative stress accounts for differential immunosuppression. In this study, we investigated whether corticosterone differentially affects the constitutive innate and humoral acquired immunity. We used barn owl nestlings, implanting 50% with a corticosterone-releasing pellet and the other 50% with a placebo pellet. To measure the effect on humoral immunity we vaccinated 50% of the corticosterone-nestlings and 50% of the placebo-nestlings with the antigens 'Tetravac' and the other 50% were injected with PBS. To assess the costs of elevated corticosterone, we measured body mass and resistance to oxidative stress. Administration of corticosterone increased corticosterone levels whereas vaccination induced the production of antibodies. Corticosterone reduced the production of antibodies, but it did not significantly affect the constitutive innate immunity. Corticosterone reduced body growth and resistance to oxidative stress. Under stressful conditions barn owl nestlings seem to keep the constitutive innate immunity, whereas elevated corticosterone levels negatively affected inducible immune responses. We found evidence that mounting a humoral immune reaction is not costly in terms of growth, but reduces the resistance to oxidative stress independently of corticosterone administration. We suggest that humoral immunity is suppressed because the risk of immunopathologies may be disproportionately high when mounting an antibody response under stressful situations.

Discovery of amino acid variants in the human glucose-dependent insulinotropic polypeptide (GIP) receptor: the impact on the pancreatic beta cell responses and functional expression studies in Chinese hamster fibroblast cells.

The two incretins, glucose-dependent insulinotropic polypeptide (GIP) and glucagon-like peptide-1 (GLP-1), are insulinotropic factors released from the small intestine to the blood stream in response to oral glucose ingestion. The insulinotropic effect of GLP-1 is maintained in patients with Type II (non-insulin-dependent) diabetes mellitus, whereas, for unknown reasons, the effect of GIP is diminished or lacking. We defined the exon-intron boundaries of the human GIP receptor, made a mutational analysis of the gene and identified two amino acid substitutions, A207 V and E354Q. In an association study of 227 Caucasian Type II diabetic patients and 224 matched glucose tolerant control subjects, the allelic frequency of the A207 V polymorphism was 1.1% in Type II diabetic patients and 0.7% in control subjects (p = 0.48), whereas the allelic frequency of the codon 354 polymorphism was 24.9% in Type II diabetic patients versus 23.2% in control subjects. Interestingly, the glucose tolerant subjects (6% of the population) who were homozygous for the codon 354 variant had on average a 14% decrease in fasting serum C-peptide concentration (p = 0.01) and an 11% decrease in the same variable 30 min after an oral glucose load (p = 0.03) compared with subjects with the wild-type receptor. Investigation of the function of the two GIP receptor variants in Chinese hamster fibroblasts showed, however, that the GIP-induced cAMP formation and the binding of GIP to cells expressing the variant receptors were not different from the findings in cells expressing the wildtype GIP receptor. In conclusion, amino acid variants in the GIP receptor are not associated with random Type II diabetes in patients of Danish Caucasian origin or with altered GIP binding and GIP-induced cAMP production when stably transfected in Chinese hamster fibroblasts. The finding of an association between homozygosity for the codon 354 variant and reduced fasting and post oral glucose tolerance test (OGTT) serum C-peptide concentrations, however, calls for further investigations and could suggest that GIP even in the fasting state regulates the beta-cell secretory response.

Glucose and leptin induce apoptosis in human beta-cells and impair glucose-stimulated insulin secretion through activation of c-Jun N-terminal kinases.

c-Jun N-terminal kinases (SAPK/JNKs) are activated by inflammatory cytokines, and JNK signaling is involved in insulin resistance and beta-cell secretory function and survival. Chronic high glucose concentrations and leptin induce interleukin-1beta (IL-1beta) secretion from pancreatic islets, an event that is possibly causal in promoting beta-cell dysfunction and death. The present study provides evidence that chronically elevated concentrations of leptin and glucose induce beta-cell apoptosis through activation of the JNK pathway in human islets and in insulinoma (INS 832/13) cells. JNK inhibition by the dominant inhibitor JNK-binding domain of IB1/JIP-1 (JNKi) reduced JNK activity and apoptosis induced by leptin and glucose. Exposure of human islets to leptin and high glucose concentrations leads to a decrease of glucose-induced insulin secretion, which was partly restored by JNKi. We detected an interplay between the JNK cascade and the caspase 1/IL-1beta-converting enzyme in human islets. The caspase 1 gene, which contains a potential activating protein-1 binding site, was up-regulated in pancreatic sections and in isolated islets from type 2 diabetic patients. Similarly, cultured human islets exposed to high glucose- and leptin-induced caspase 1 and JNK inhibition prevented this up-regulation. Therefore, JNK inhibition may protect beta-cells from the deleterious effects of high glucose and leptin in diabetes.

Exercise aggravates cardiovascular risks and mortality in rats with disrupted nitric oxide pathway and treated with recombinant human erythropoietin.

Chronic administration of recombinant human erythropoietin (rHuEPO) can generate serious cardiovascular side effects such as arterial hypertension (HTA) in clinical and sport fields. It is hypothesized that nitric oxide (NO) can protect from noxious cardiovascular effects induced by chronic administration of rHuEPO. On this base, we studied the cardiovascular effects of chronic administration of rHuEPO in exercise-trained rats treated with an inhibitor of NO synthesis (L-NAME). Rats were treated or not with rHuEPO and/or L-NAME during 6 weeks. During the same period, rats were subjected to treadmill exercise. The blood pressure was measured weekly. Endothelial function of isolated aorta and small mesenteric arteries were studied and the morphology of the latter was investigated. L-NAME induced hypertension (197 ± 6 mmHg, at the end of the protocol). Exercise prevented the rise in blood pressure induced by L-NAME (170 ± 5 mmHg). However, exercise-trained rats treated with both rHuEPO and L-NAME developed severe hypertension (228 ± 9 mmHg). Furthermore, in these exercise-trained rats treated with rHuEPO/L-NAME, the acetylcholine-induced relaxation was markedly impaired in isolated aorta (60% of maximal relaxation) and small mesenteric arteries (53%). L-NAME hypertension induced an internal remodeling of small mesenteric arteries that was not modified by exercise, rHuEPO or both. Vascular ET-1 production was not increased in rHuEPO/L-NAME/training hypertensive rats. Furthermore, we observed that rHuEPO/L-NAME/training hypertensive rats died during the exercise or the recovery period (mortality 51%). Our findings suggest that the use of rHuEPO in sport, in order to improve physical performance, represents a high and fatal risk factor, especially with pre-existing cardiovascular risk.