Early Increase in Blood-Brain Barrier Permeability in a Murine Model Exposed to Fifteen Days of Intermittent Hypoxia.

Obstructive sleep apnea (OSA) is characterized by intermittent repeated episodes of hypoxia-reoxygenation. OSA is associated with cerebrovascular consequences. An enhanced blood-brain barrier (BBB) permeability has been proposed as a marker of those disorders. We studied in mice the effects of 1 day and 15 days intermittent hypoxia (IH) exposure on BBB function. We focused on the dorsal part of the hippocampus and attempted to identify the molecular mechanisms by combining in vivo BBB permeability (Evans blue tests) and mRNA expression of several junction proteins (zona occludens (ZO-1,2,3), VE-cadherin, claudins (1,5,12), cingulin) and of aquaporins (1,4,9) on hippocampal brain tissues. After 15 days of IH exposure we observed an increase in BBB permeability, associated with increased mRNA expressions of claudins 1 and 12, aquaporins 1 and 9. IH seemed to increase early for claudin-1 mRNA expression as it doubled with 1 day of exposure and returned near to its base level after 15 days. Claudin-1 overexpression may represent an immediate response to IH exposure. Then, after 15 days of exposure, an increase in functional BBB permeability was associated with enhanced expression of aquaporin. These BBB alterations are possibly associated with a vasogenic oedema that may affect brain functions and accelerate neurodegenerative processes.

Intravenous injection of a novel viral immunotherapy encoding human interleukin-7 in nonhuman primates is safe and increases absolute lymphocyte count.

Persistence of an immunosuppression, affecting both the innate and adaptive arms of the immune system, plays a role in sepsis patients' morbidity and late mortality pointing to the need for broad and effective immune interventions. MVA-hIL-7-Fc is a non-replicative recombinant Modified Vaccinia virus Ankara encoding the human interleukin-7 fused to human IgG2 Fc fragment. We have shown in murine sepsis models the capacity of this new virotherapy to stimulate both arms of the immune system and increase survival. Herein, an exploratory study in nonhuman primates was performed following a single intravenous injection of the MVA-hIL-7-Fc used at the clinical dose to assess its safety and biological activities. Four cynomolgus macaques were followed for 3 weeks post-injection (p.i), without observed acute adverse reactions. Circulating hIL-7-Fc was detected during the first 3-5 days p.i with a detection peaking at 12 h p.i. IL-7 receptor engagement and downstream signal transduction were detected in T cells demonstrating functionality of the expressed IL-7. Expansion of blood lymphocytes, mainly CD4 and CD8 naïve and central memory T cells, was observed on day 7 p.i. together with a transient increase of Ki67 expression on T lymphocytes. In addition, we observed an increase in circulating B and NK cells as well as monocytes were albeit with different kinetics and levels. This study indicates that a vectorized IL-7-Fc, injected by intravenous route at a relevant clinical dose in a large animal model, is active without adverse reactions supporting the clinical development of this novel virotherapy for treatment of sepsis patients.

Viral Delivery of IL-7 Is a Potent Immunotherapy Stimulating Innate and Adaptive Immunity and Confers Survival in Sepsis Models.

Persistence of an immunosuppressive state plays a role in septic patient morbidity and late mortality. Both innate and adaptive pathways are impaired, pointing toward the need for immune interventions targeting both arms of the immune system. We developed a virotherapy using the nonpropagative modified vaccinia virus Ankara (MVA), which harbors the intrinsic capacity to stimulate innate immunity, to deliver IL-7, a potent activator of adaptive immunity. The rMVA-human IL-7 (hIL-7)-Fc encoding the hIL-7 fused to the human IgG2-Fc was engineered and shown to express a dimeric, glycosylated, and biologically active cytokine. Following a single i.v. injection in naive mice, the MVA-hIL-7-Fc increased the number of total and activated B, T, and NK cells but also myeloid subpopulations (Ly6Chigh, Ly6Cint, and Ly6Cneg cells) in both lung and spleen. It triggered differentiation of T cells in central memory, effector memory, and acute effector phenotypes and enhanced polyfunctionality of T cells, notably the number of IFN-γ-producing cells. The MVA vector contributed significantly to immune cell activation, particularly of NK cells. The MVA-hIL-7-Fc conferred a significant survival advantage in the cecal ligation and puncture (CLP) and Candida albicans sepsis models. It significantly increased cell numbers and activation in both spleen and lung of CLP mice. Comparatively, in naive and CLP mice, the rhIL-7-Fc soluble counterpart overall induced less vigorous, shorter lasting, and narrower immune activities than did the MVA-hIL-7-Fc and favored TNF-α-producing cells. The MVA-hIL-7-Fc represents a novel class of immunotherapeutic with clinical potential for treatment of septic patients.

Chronic Intermittent Hypoxia Alters Rat Ophthalmic Artery Reactivity Through Oxidative Stress, Endothelin and Endothelium-Derived Hyperpolarizing Pathways.

Purpose: Obstructive sleep apnea recently has been associated with a higher frequency of ischemic optic neuropathies. Intermittent hypoxia (IH) has been proposed as a major component of obstructive sleep apnea cardiovascular consequences. However, there currently are no pathophysiologic data regarding the effect of IH on the ocular vascular system. Thus, we assessed the impact of chronic IH exposure on the morphology and vascular reactivity of the rat ophthalmic artery (OA). Methods: Rats were exposed to 14 days of IH or normoxia (NX). Ophthalmic artery reactivity was studied using wire myography in rats treated or not with tempol (1 mM/day). Expression of endothelin-1 (ET-1) and its receptors, and of the three nitric oxide synthase (NOS) isoform genes was quantified using quantitative polymerase chain reaction (qPCR) in the retina and optic nerve. Structural alterations (optical and electron microscopy) and superoxide anion production were studied in OA sections. Results: Superoxide ion expression in the OA wall was increased by 23% after IH exposure. Ophthalmic artery contractile response to 3.10-8 M ET-1 was increased by 18.6% and nitric oxide-mediated relaxation was significantly delayed in IH compared to NX rats. In the absence of nitric oxide, cytochrome P450 blockade increased relaxation to acetylcholine in IH rats and delayed it in NX rats. Tempol treatment abolished the IH-induced changes in OA reactivity. Conclusions: These results strongly suggest that chronic IH induces oxidative stress in the rat OA, associated with endothelial dysfunction through alterations of nitric oxide and endothelium-derived hyperpolarising factors (EDHF) pathways.

Atorvastatin protects against deleterious cardiovascular consequences induced by chronic intermittent hypoxia.

Chronic intermittent hypoxia (IH), a major component of obstructive sleep apnea (OSA), contributes to the high risk of cardiovascular morbidity. We have previously demonstrated that IH-induced oxidative stress is involved in the hypertension and in the hypersensitivity to myocardial infarction. However, the mechanisms underlying these cardiovascular alterations are still unclear, as well as the role of potential protective treatment. Atorvastatin has pleiotropic actions, including increasing nitric oxide (NO) bioavailability and reducing inflammation and oxidative damage. The aim of this study was to evaluate the beneficial effect of a two time course of this treatment against the deleterious cardiovascular consequences of IH. Rats were divided into two groups subjected to chronic IH or normoxic (N) exposure. IH consisted of repetitive one-minute cycles (with only 30 s of a 5% inspired O2 fraction) and was applied for eight hours during daytime, for 14 (simultaneous protocol) or 28 d (delayed protocol). Atorvastatin (10 mg/kg/ d) or its vehicle was administered during the 14 d simultaneous protocol or the last 14 d of the delayed protocol. For both protocols, systolic arterial pressure was significantly increased by 14 d IH exposure. Atorvastatin prevented this deleterious effect in the simultaneous protocol. Carotid artery compliance and endothelial function were significantly altered after 28 d but not after 14 d of IH exposure. Delayed atorvastatin administration preserved these vascular parameters. IH also increased hypersensitivity to myocardial infarction after 14 d exposure, and atorvastatin abolished this deleterious effect. IH also enhanced cardiac NADPH expression and decreased aortic superoxide dismutase activity after 14 d exposure. Atorvastatin significantly restored these activities. In conclusion, whereas IH rapidly increased blood pressure, myocardial infarction hypersensitivity and oxidative stress, compliance, endothelial function and the structural wall of the carotid artery were only altered after a longer IH exposure. Atorvastatin prevented all these deleterious cardiovascular effects, leading to a potentially novel pharmacological therapeutic strategy for OSA syndrome.

Sex effect in mouse and human prion disease.

Sex effect on the incubation period of variant Creutzfeldt-Jakob disease (vCJD) disease in human and ME-7 murine models was investigated. In the 167 vCJD cases reported in the United Kingdom as of January 2009, age at onset was significantly lower in female patients (by 2 years) than in male patients after stratification on birth cohort. In C57/Bl6N mice infected with ME-7 scrapie strain, incubation was shorter in female than in male mice. The incubation period increased in castrated male mice after intraperitoneal infection but not after intracerebral inoculation. In the absence of androgen receptors, the incubation period for prion disease increased after intraperitoneal inoculation. In ovariectomized or estrogen receptor alpha-defective female mice, no effect was observed on the incubation period of mouse prion disease. These results show that androgens influence the prion diseases incubation period after inoculation at a peripheral site.