Bioinspired Scaffold Action Under the Extreme Physiological Conditions of Simulated Space Flights: Osteogenesis Enhancing Under Microgravity.

Prolonged exposure to microgravity (MG) during long-duration space flights is known to induce severe dysregulation of osteoblast functions connected to a significant bone loss, similar to the condition induced by osteoporosis. Hence, we here present MG as a promising model to challenge the effectiveness of new scaffolds designed for bone regeneration in counteracting bone loss. To this end, we carried out an integrative study aimed to evaluate, in the extreme condition of Random Positioning Machine-simulated MG, the osteoinductive potential of nanocrystalline magnesium-doped hydroxyapatite/type I collagen composite scaffold (MHA/Coll), that we previously demonstrated to be an excellent tool for bone tissue engineering.

Band 3 Erythrocyte Membrane Protein Acts as Redox Stress Sensor Leading to Its Phosphorylation by p (72) Syk.

In erythrocytes, the regulation of the redox sensitive Tyr phosphorylation of band 3 and its functions are still partially defined. A role of band 3 oxidation in regulating its own phosphorylation has been previously suggested. The current study provides evidences to support this hypothesis: (i) in intact erythrocytes, at 2 mM concentration of GSH, band 3 oxidation, and phosphorylation, Syk translocation to the membrane and Syk phosphorylation responded to the same micromolar concentrations of oxidants showing identical temporal variations; (ii) the Cys residues located in the band 3 cytoplasmic domain are 20-fold more reactive than GSH; (iii) disulfide linked band 3 cytoplasmic domain docks Syk kinase; (iv) protein Tyr phosphatases are poorly inhibited at oxidant concentrations leading to massive band 3 oxidation and phosphorylation.

Signal transduction in primary human T lymphocytes in altered gravity during parabolic flight and clinostat experiments.

Background/aims: Several limiting factors for human health and performance in microgravity have been clearly identified arising from the immune system, and substantial research activities are required in order to provide the basic information for appropriate integrated risk management. The gravity-sensitive nature of cells of the immune system renders them an ideal biological model in search for general gravity-sensitive mechanisms and to understand how the architecture and function of human cells is related to the gravitational force and therefore adapted to life on Earth.

Methods: We investigated the influence of altered gravity in parabolic flight and 2D clinostat experiments on key proteins of activation and signaling in primary T lymphocytes.

T cell tyrosine phosphorylation response to transient redox stress.

Reactive Oxygen Species (ROS) are crucial to multiple biological processes involved in the pathophysiology of inflammation, and are also involved in redox signaling responses. Although previous reports have described an association between oxidative events and the modulation of innate immunity, a role for redox signaling in T cell mediated adaptive immunity has not been described yet. This work aims at assessing if T cells can sense redox stress through protein sulfhydryl oxidation and respond with tyrosine phosphorylation changes.

Immunomodulatory properties of carbon nanotubes are able to compensate immune function dysregulation caused by microgravity conditions.

Spaceflights lead to dysregulation of the immune cell functionality affecting the expression of activation markers and cytokine production. Short oxidized multi-walled carbon nanotubes functionalized by 1,3-dipolar cycloaddition have been reported to activate immune cells. In this Communication we have performed surface marker assays and multiplex ELISA on primary monocytes and T cells under microgravity.

Signal transduction in primary human T lymphocytes in altered gravity - results of the MASER-12 suborbital space flight mission.

We investigated the influence of altered gravity on key proteins of T cell activation during the MASER-12 ballistic suborbital rocket mission of the European Space Agency (ESA) and the Swedish Space Cooperation (SSC) at ESRANGE Space Center (Kiruna, Sweden). We quantified components of the T cell receptor, the membrane proximal signaling, MAPK-signaling, IL-2R, histone modifications and the cytoskeleton in non-activated and in ConA/CD28-activated primary human T lymphocytes. The hypergravity phase during the launch resulted in a downregulation of the IL-2 and CD3 receptor and reduction of tyrosine phosphorylation, p44/42-MAPK phosphorylation and histone H3 acetylation, whereas LAT phosphorylation was increased.

Effect of heterozygous beta thalassemia on the phosphorylative response to Plasmodium falciparum infection.

Malaria parasites interact with the host cell membrane inserting new proteins and inducing oxidative and phosphorylative changes of erythrocyte proteins. In the present report we monitored the time dependent oxidative and phosphorylative modifications induced by parasites in heterozygous beta thalassemia (Het-βThal). Het-βThal causes mild anemia and is known to determine a pro-oxidant milieu and a protective effect against severe malaria.

The Rel/NF-κB pathway and transcription of immediate early genes in T cell activation are inhibited by microgravity.

This study tested the hypothesis that transcription of immediate early genes is inhibited in T cells activated in μg. Immunosuppression during spaceflight is a major barrier to safe, long-term human space habitation and travel. The goals of these experiments were to prove that μg was the cause of impaired T cell activation during spaceflight, as well as understand the mechanisms controlling early T cell activation.

5-Lipoxygenase-dependent apoptosis of human lymphocytes in the International Space Station: data from the ROALD experiment.

The functional adaptation of the immune system to the surrounding environment is also a fundamental issue in space. It has been suggested that a decreased number of lymphocytes might be a cause of immunosuppression, possibly due to the induction of apoptosis. Early activation of 5-lipoxygenase (5-LOX) might play a central role in the initiation of the apoptotic program.

New antimalarial indolone-N-oxides, generating radical species, destabilize the host cell membrane at early stages of Plasmodium falciparum growth: role of band 3 tyrosine phosphorylation.

Although indolone-N-oxide (INODs) genereting long-lived radicals possess antiplasmodial activity in the low-nanomolar range, little is known about their mechanism of action. To explore the molecular basis of INOD activity, we screened for changes in INOD-treated malaria-infected erythrocytes (Pf-RBCs) using a proteomics approach. At early parasite maturation stages, treatment with INODs at their IC(50) concentrations induced a marked tyrosine phosphorylation of the erythrocyte membrane protein band 3, whereas no effect was observed in control RBCs.

Space flight affects motility and cytoskeletal structures in human monocyte cell line J-111.

Certain functions of immune cells in returning astronauts are known to be altered. A dramatic depression of the mitogenic in vitro activation of human lymphocytes was observed in low gravity. T-cell activation requires the interaction of different type of immune cells as T-lymphocytes and monocytes.

Irreversible AE1 tyrosine phosphorylation leads to membrane vesiculation in G6PD deficient red cells.

Background: While G6PD deficiency is one of the major causes of acute hemolytic anemia, the membrane changes leading to red cell lysis have not been extensively studied. New findings concerning the mechanisms of G6PD deficient red cell destruction may facilitate our understanding of the large individual variations in susceptibility to pro-oxidant compounds and aid the prediction of the hemolytic activity of new drugs.

Methodology/principal Findings: Our results show that treatment of G6PD deficient red cells with diamide (0.

Modification of proteins secreted by endothelial cells during modeled low gravity exposure.

The exposure of the human body to microgravity, conditions that occurs during space flights, causes significant changes in the cardiovascular system. Many cell types have been involved in these changes, and the endothelium seems to play a major role. In endothelial cells (EC), it has been shown that modeled low gravity impairs nitric oxide synthesis, cell adhesion, extracellular matrix composition, cytoskeleton organization, cytokines, and growth factors secretion.

Analysis of changes in tyrosine and serine phosphorylation of red cell membrane proteins induced by P. falciparum growth.

Phosphorylation of erythrocyte membrane proteins has been previously documented following infection and intracellular growth of the malarial parasite, Plasmodium falciparum in red cells. Much of this data dealt with phosphorylation of serine residues. In this study, we report detailed characterization of phosphorylation of serine and tyrosine residues of red cell membrane proteins following infection by P falciparum.

Involvement of adenosine A1 and A2A receptors in (-)-linalool-induced antinociception.

In recent studies performed in our laboratory we have shown that acute administration of (-)-linalool, the natural occurring enantiomer in essential oils, possesses anti-inflammatory, antihyperalgesic and antinociceptive effects in different animal models. The antihyperalgesic and antinociceptive effects of (-)-linalool have been ascribed to its capacity in stimulating the opioidergic, cholinergic and dopaminergic systems, as well as to its interaction with K+ channels, or to its local anaesthetic activity and/or to the negative modulation of glutamate transmission. Activation of A1 or A2A receptors has been shown to induce antinociceptive effects, and the possible involvement of adenosine in (-)-linalool antinociceptive effect, has not been elucidated yet.

Microgravity alters basal and insulin-mediated metabolic activity of normal and neoplastic cells.

In this paper we report the behaviour of normal vascular smooth muscle cells and transformed breast cancer cells under normal versus simulated microgravity conditions by comparing cell proliferation, Glucose transport, Methionine uptake and protein synthesis. Modeled microgravity profoundly affects cell growth (especially in normal cells) and Glucose or Methionine metabolism (although to different extent in the two cell lines). Since both cells own responsive insulin receptors, the comparison was extended to insulin-stimulated versus unstimulated conditions.

Exposure of human lymphocytes and lymphoblastoid cells to simulated microgravity strongly affects energy metabolism and DNA repair.

Exposure of freshly drawn lymphocytes and lymphoblastoid cells (LB and COR3) to simulated microgravity decreased the intracellular ATP concentration to 50%-40% of the value found in normal growth conditions. The decrease was reversible although recovery to normal values occurred only slowly both in lymphocytes and in lymphoblastoid cells. Poly(ADP-ribose) polymerase (PARP ) activity was increased indicating that cells exposed to conditions of reduced gravitation experience stress.

Effects of (-)-linalool in the acute hyperalgesia induced by carrageenan, L-glutamate and prostaglandin E2.

A series of studies performed in our laboratory have shown that (-)-linalool, the natural occurring enantiomer in essential oils, possesses anti-inflammatory and antinociceptive effects in different animal models. The antinociceptive effect of (-)-linalool has been ascribed to the stimulation of the cholinergic, opioidergic and dopaminergic systems, to its local anesthetic activity and to the blockade of N-methyl-D-aspartate (NMDA) receptors. In this study, we investigated the effect of systemic administration of (-)-linalool in the paw withdrawal test in rats, a model of thermal hyperalgesia induced by monolateral subplantar injection of carrageenan, L-glutamate or prostaglandin E(2).

Profile of spinal and supra-spinal antinociception of (-)-linalool.

We previously reported that administration of (-)-linalool, the naturally occurring enantiomer in essential oils, induced a significant reduction in carrageenin-induced oedema and in acetic acid-induced writhing. The latter effect was completely antagonised by the muscarinic receptor antagonist atropine and by the opioid receptor antagonist naloxone. To further characterise the antinociceptive profile of (-)-linalool, we studied its effect in the hot plate and the formalin in tests.

Creating conditions similar to those that occur during exposure of cells to microgravity induces apoptosis in human lymphocytes by 5-lipoxygenase-mediated mitochondrial uncoupling and cytochrome c release.

Creating conditions similar to those that occur during exposure of cells to microgravity induced a sixfold increase of apoptotic bodies and DNA fragments in human lymphocytes, paralleled by an early (within 2 h) fourfold increase in 5-lipoxygenase (5-LOX) activity and a fivefold decrease in mitochondrial membrane potential and increase in cytochrome c release (within 4 and 8 h, respectively). Similar membrane potential and cytochrome c release were observed in isolated mitochondria treated with physiological amounts of 5-LOX and were enhanced by creating conditions similar to those that occur during exposure of cells to microgravity. 5-LOX inhibitors, 5,8,11,14-eicosatetraynoic acid and caffeic acid, completely prevented apoptosis, whereas the phospholipase A(2) inhibitor methyl-arachidonoyl fluorophosphonate and the 5-LOX activating protein inhibitor MK886 reduced it to 65-70%.

(-)-Linalool produces antinociception in two experimental models of pain.

Linalool is a monoterpene compound commonly found as a major component of the essential oils of several aromatic plant species, many of which are used in traditional medical systems as analgesic and anti-inflammatory remedies. We previously reported that (-)-linalool, the natural occurring enantiomer, plays a major role in the anti-inflammatory activity displayed by different essential oils, suggesting that linalool-producing species are potentially anti-inflammatory agents. In this study, the antinociceptive activity of (-)-linalool was examined in two different pain models in mice: the acetic acid-induced writhing response, a model of inflammatory pain, and the hot plate test, a model of supraspinal analgesia.