Compatibility of Nucleobases Containing Pt(II) Complexes with Red Blood Cells for Possible Drug Delivery Applications.

The therapeutic advantages of some platinum complexes as major anticancer chemotherapeutic agents and of nucleoside analogue-based compounds as essential antiviral/antitumor drugs are widely recognized. Red blood cells (RBCs) offer a potential new strategy for the targeted release of therapeutic agents due to their biocompatibility, which can protect loaded drugs from inactivation in the blood, thus improving biodistribution. In this study, we evaluated the feasibility of loading model nucleobase-containing Pt(II) complexes into human RBCs that were highly stabilized by four N-donors and susceptible to further modification for possible antitumor/antiviral applications. Specifically, platinum-based nucleoside derivatives [PtII(dien)(N7-Guo)]2+, [PtII(dien)(N7-dGuo)]2+, and [PtII(dien)(N7-dGTP)] (dien = diethylenetriamine; Guo = guanosine; dGuo = 2'-deoxy-guanosine; dGTP = 5'-(2'-deoxy)-guanosine-triphosphate) were investigated. These Pt(II) complexes were demonstrated to be stable species suitable for incorporation into RBCs. This result opens avenues for the possible incorporation of other metalated nucleobases analogues, with potential antitumor and/or antiviral activity, into RBCs.

Tunneling nanotubes and mesenchymal stem cells: New insights into the role of melatonin in neuronal recovery.

Efficient cell-to-cell communication is essential for tissue development, homeostasis, and the maintenance of cellular functions after injury. Tunneling nanotubes (TNTs) have emerged as a new important method of cell-to-cell communication. TNTs are primarily established between stressed and unstressed cells and can transport a variety of cellular components. Mitochondria are important trafficked entities through TNTs. Transcellular mitochondria transfer permits the incorporation of healthy mitochondria into the endogenous network of recipient cells, changing the bioenergetic profile and other functional properties of the recipient and may allow the recipient cells to recuperate from apoptotic processes and return to a normal operating state. Mesenchymal cells (MSCs) can form TNTs and transfer mitochondria and other constituents to target cells. This occurs under both physiological and pathological conditions, leading to changes in cellular energy metabolism and functions. This review summarizes the newly described capacity of melatonin to improve mitochondrial fusion/fission dynamics and promote TNT formation. This new evidence suggests that melatonin's protective effects could be attributed to its ability to prevent mitochondrial damage in injured cells, reduce senescence, and promote anastasis, a natural cell recovery phenomenon that rescues cells from the brink of death. The modulation of these new routes of intercellular communication by melatonin could play a key role in increasing the therapeutic potential of MSCs.

Extracellular Vesicles from Campylobacter jejuni CDT-Treated Caco-2 Cells Inhibit Proliferation of Tumour Intestinal Caco-2 Cells and Myeloid U937 Cells: Detailing the Global Cell Response for Potential Application in Anti-Tumour Strategies.

Cytolethal distending toxin (CDT) is produced by a range of Gram-negative pathogenic bacteria such as Campylobacter jejuni. CDT represents an important virulence factor that is a heterotrimeric complex composed of CdtA, CdtB, and CdtC. CdtA and CdtC constitute regulatory subunits whilst CdtB acts as the catalytic subunit exhibiting phosphatase and DNase activities, resulting in cell cycle arrest and cell death. Extracellular vesicle (EV) secretion is an evolutionarily conserved process that is present throughout all kingdoms. Mammalian EVs play important roles in regular cell-to-cell communications but can also spread pathogen- and host-derived molecules during infections to alter immune responses. Here, we demonstrate that CDT targets the endo-lysosomal compartment, partially evading lysosomal degradation and exploiting unconventional secretion (EV release), which is largely involved in bacterial infections. CDT-like effects are transferred by Caco-2 cells to uninfected heterologous U937 and homologous Caco-2 cells. The journey of EVs derived from CDT-treated Caco-2 cells is associated with both intestinal and myeloid tumour cells. EV release represents the primary route of CDT dissemination, revealing an active toxin as part of the cargo. We demonstrated that bacterial toxins could represent suitable tools in cancer therapy, highlighting both the benefits and limitations. The global cell response involves a moderate induction of apoptosis and autophagic features may play a protective role against toxin-induced cell death. EVs from CDT-treated Caco-2 cells represent reliable CDT carriers, potentially suitable in colorectal cancer treatments. Our data present a potential bacterial-related biotherapeutic supporting a multidrug anticancer protocol.

Synthesis and biological characterization of a new fluorescent probe for vesicular trafficking based on polyazamacrocycle derivative.

The fluorescent probes represent an important tool in the biological study, in fact characterization of cellular structures and organelles are an important tool-target for understanding the mechanisms regulating most biological processes. Recently, a series of polyamino-macrocycles based on 1,4,7,10-tetraazacyclododecane was synthesized, bearing one or two NBD units (AJ2NBD·4HCl) useful as sensors for metal cations and halides able to target and to detect apolar environment, as lipid membranes. In this paper, we firstly illustrate the chemical synthesis of the AJ2NBD probe, its electronic absorption spectra and its behavior regarding pH of the environment. Lack of any cellular toxicity and an efficient labelling on fresh, living cells was demonstrated, allowing the use of AJ2NBD in biological studies. In particular, this green fluorescent probe may represent a potential dye for the compartments involved in the endosomal/autophagic pathway. This research's field should benefit from the use of AJ2NBD as a vesicular tracer, however, to ensure the precise nature of vesicles/vacuoles traced by this new probe, other more specific tests are needed.

Serum changes in sTWEAK and its scavenger receptor sCD163 in ultramarathon athletes running the 24-h race.

In the present investigation, the serum changes of sTWEAK levels, a multifunctional cytokine involved in tissue response to acute injury and inflammation, and of its scavenger receptor sCD163, were monitored for the first time in ultramarathon athletes running the 24-h competition, an extremely demanding race in terms of muscular and physiological exertion. To this aim, venous blood samples were collected from each participant (n = 22, M = 12, F = 10) both before and immediately after the 24-h running. Other than sTWEAK and sCD163, the common serum biomarkers of inflammation (namely CRP and IL-6) and tissue injury (such as CPK, LDH, CPK-MB, troponin-I, and NT-proBNP) were evaluated. All parameters were within the reference ranges at baseline, indicating no alterations of the normal physiological processes before the competition; on the contrary, most biomarkers of tissue damage and inflammation strongly increased after the ultramarathon race. Interestingly, a significant decrement of sTWEAK levels associated with an increment of its scavenger receptor sCD163 was observed at post-race. Positive relationships were evidenced between IL-6 and sCD163 levels and the markers of cardiac damage troponin-I and NT-proBNP. On the contrary, sTWEAK showed an inverse correlation with IL-6 and NT-proBNP. This study opens the way to further investigations aimed at clarifying the role of TWEAK pathway during the prolonged ultraendurance activity, paying particular attention to the link of IL-6, CD163 and TWEAK with the cardiac function.

Melatonin reshapes the mitochondrial network and promotes intercellular mitochondrial transfer via tunneling nanotubes after ischemic-like injury in hippocampal HT22 cells.

Mitochondrial dysfunction is considered one of the hallmarks of ischemia/reperfusion injury. Mitochondria are plastic organelles that undergo continuous biogenesis, fusion, and fission. They can be transferred between cells through tunneling nanotubes (TNTs), dynamic structures that allow the exchange of proteins, soluble molecules, and organelles. Maintaining mitochondrial dynamics is crucial to cell function and survival. The present study aimed to assess the effects of melatonin on mitochondrial dynamics, TNT formation, and mitochondria transfer in HT22 cells exposed to oxygen/glucose deprivation followed by reoxygenation (OGD/R). The results showed that melatonin treatment during the reoxygenation phase reduced mitochondrial reactive oxygen species (ROS) production, improved cell viability, and increased the expression of PGC1α and SIRT3. Melatonin also preserved the expression of the membrane translocase proteins TOM20 and TIM23, and of the matrix protein HSP60, which are involved in mitochondrial biogenesis. Moreover, it promoted mitochondrial fusion and enhanced the expression of MFN2 and OPA1. Remarkably, melatonin also fostered mitochondrial transfer between injured HT22 cells through TNT connections. These results provide new insights into the effect of melatonin on mitochondrial network reshaping and cell survival. Fostering TNTs formation represents a novel mechanism mediating the protective effect of melatonin in ischemia/reperfusion injury.

Rapamycin Re-Directs Lysosome Network, Stimulates ER-Remodeling, Involving Membrane CD317 and Affecting Exocytosis, in Campylobacter Jejuni-Lysate-Infected U937 Cells.

The Gram-negative Campylobacter jejuni is a major cause of foodborne gastroenteritis in humans worldwide. The cytotoxic effects of Campylobacter have been mainly ascribed to the actions of the cytolethal distending toxin (CDT): it is mandatory to put in evidence risk factors for sequela development, such as reactive arthritis (ReA) and Guillain-Barré syndrome (GBS). Several researches are directed to managing symptom severity and the possible onset of sequelae. We found for the first time that rapamycin (RM) is able to largely inhibit the action of C. jejuni lysate CDT in U937 cells, and to partially avoid the activation of specific sub-lethal effects. In fact, we observed that the ability of this drug to redirect lysosomal compartment, stimulate ER-remodeling (highlighted by ER-lysosome and ER-mitochondria contacts), protect mitochondria network, and downregulate CD317/tetherin, is an important component of membrane microdomains. In particular, lysosomes are involved in the process of the reduction of intoxication, until the final step of lysosome exocytosis. Our results indicate that rapamycin confers protection against C. jejuni bacterial lysate insults to myeloid cells.

New insights into the trophic and cytoprotective effects of creatine in in vitro and in vivo models of cell maturation.

A growing body of scientific reports indicates that the role of creatine (Cr) in cellular biochemistry and physiology goes beyond its contribution to cell energy. Indeed Cr has been shown to exert multiple effects promoting a wide range of physiological responses in vitro as well as in vivo. Included in these, Cr promotes in vitro neuron and muscle cell differentiation, viability and survival under normal or adverse conditions; anabolic, protective and pro-differentiative effects have also been observed in vivo. For example Cr has been shown to accelerate in vitro differentiation of cultured C2C12 myoblasts into myotubes, where it also induces a slight but significant hypertrophic effect as compared to unsupplemented cultures; Cr also prevents the anti-differentiation effects caused by oxidative stress in the same cells. In trained adults, Cr increases the mRNA expression of relevant myogemic factors, protein synthesis, muscle strength and size, in cooperation with physical exercise. As to neurons and central nervous system, Cr favors the electrophysiological maturation of chick neuroblasts in vitro and protects them from oxidative stress-caused killing; similarly, Cr promotes the survival and differentiation of GABA-ergic neurons in fetal spinal cord cultures in vitro; in vivo, maternal Cr supplementation promotes the morpho-functional development of hippocampal neurons in rat offsprings. This article, which presents also some new experimental data, focuses on the trophic, pro-survival and pro-differentiation effects of Cr and examines the ensuing preventive and therapeutic potential in pathological muscle and brain conditions.

Melatonin regulates mesenchymal stem cell differentiation: a review.

Among the numerous functions of melatonin, the control of survival and differentiation of mesenchymal stem cells (MSCs) has been recently proposed. MSCs are a heterogeneous population of multipotent elements resident in tissues such as bone marrow, muscle, and adipose tissue, which are primarily involved in developmental and regeneration processes, gaining thus increasing interest for tissue repair and restoration therapeutic protocols. Receptor-dependent and receptor-independent responses to melatonin are suggested to occur in these cells. These involve antioxidant or redox-dependent functions of this indolamine as well as secondary effects resulting from autocrine and paracrine responses. Inflammatory cytokines and adipokines, proangiogenic/mitogenic stimuli, and other mediators that influence the differentiation processes may affect the survival and functional integrity of these mesenchymal precursor cells. In this scenario, melatonin seems to regulate signaling pathways that drive commitment and differentiation of MSC into osteogenic, chondrogenic, adipogenic, or myogenic lineages. Common pathways suggested to be involved as master regulators of these processes are the Wnt/ß-catenin pathway, the MAPKs and the, TGF-ß signaling. In this respect melatonin emerges a novel and potential modulator of MSC lineage commitment and adipogenic differentiation. These and other aspects of the physiological and pharmacological effects of melatonin as regulator of MSC are discussed in this review.

Pharmacological doses of melatonin induce alterations in mitochondrial mass and potential, bcl-2 levels and K+ currents in UVB-exposed U937 cells.

Apoptosis is observed in 'actively' dying cells after the exposure to cell stressors such as ultraviolet light irradiation. Since melatonin has been proposed to act under stressful conditions as cell protection factor, in this study we examined the potential of this molecule when used at pharmacological concentrations to control mitochondrial damage and apoptotic signalling of UVB irradiated U937 human leukaemic cells. Moreover, the effect of melatonin treatment on electrophysiological properties and membrane K(+) currents of irradiated U937 cells was investigated as functional aspects relevant to the anti-apoptotic role of melatonin. The general effect is associated with the restoration of mass, number and membrane potential of mitochondria, with a lower caspase activation and bcl-2 upregulation. In the presence of the caspase inhibitor ZVAD-Fmk, melatonin seems to drive UVB stressed cells to follow the mitochondrial intrinsic pathway, interfering just at the mitochondrial level. Moreover, treatment with melatonin, as well as ZVAD-Fmk, prevented the K(+) current reduction observed late following the UVB insult application, by sparing cells from death; this result also indicates that the decrease of K(+) leakage currents could represent a functional feature of apoptotic process in UV-exposed U937 cells.

New insights into the cytotoxic effects of Thymus vulgaris essential oil on the human triple-negative breast cancer cell line MDA-MB-231.

Essential oils (EOs) are natural products that have gained wide interest due to their biological activities and anticancer properties through various mechanisms. The present study aimed to test the cytotoxicity of Thymus vulgaris L. (thyme) EO of Italian origin, rich in thymol (49.6%) and p-cymene (18.8%), towards the triple-negative breast cancer cell line MDA-MB-231 and to investigate the biochemical mechanisms underlying its antitumor activity. Thyme EO reduced cancer cell viability in a dose-dependent manner after 24 h treatment, with an IC50 value equal to 75.1 ± 15.2 µg/ml; simultaneously, the inhibition of cancer cell migration and colony formation capacity was evidenced. Thyme EO antiproliferative effects were related to the induction of apoptosis as demonstrated by the increased expression of the pro-apoptotic proteins Bax, cleaved caspase-3, phospho-p53, and SMAC/Diablo and by the reduction of the anti-apoptotic proteins Bcl-2, cIAP-1, cIAP-2, HIF-1α, survivin, and XIAP. Thyme EO administration led to the early formation of intracellular ROS, followed by the increment of MDA as an index of lipid peroxidation and by the decreased expression of the antioxidant enzymes catalase and PON2. The upregulation of Nrf2 mRNA expression and the strong induction of HO-1 sustained the activation of the Nrf2 pathway by thyme EO. These data showed that the EO from Thymus vulgaris L. might inhibit the malignant phenotype of MDA-MB-231, thus suggesting potential benefits against human triple-negative breast cancer.

Melatonin, tunneling nanotubes, mesenchymal cells, and tissue regeneration.

Mesenchymal stem cells are multipotent stem cells that reside in many human tissues and organs. Mesenchymal stem cells are widely used in experimental and clinical regenerative medicine due to their capability to transdifferentiate into various lineages. However, when transplanted, they lose part of their multipotency and immunomodulatory properties, and most of them die after injection into the damaged tissue. In this review, we discuss the potential utility of melatonin in preserving mesenchymal stem cells' survival and function after transplantation. Melatonin is a pleiotropic molecule regulating critical cell functions including apoptosis, endoplasmic reticulum stress, and autophagy. Melatonin is also synthesized in the mitochondria where it reduces oxidative stress, the opening of the mitochondrial permeability transition pore and the downstream caspase activation, activates uncoupling proteins, and curtails the proinflammatory response. In addition, recent findings showed that melatonin also promotes the formation of tunneling nanotubes and the transfer of mitochondria between cells through the connecting tubules. As mitochondrial dysfunction is a primary cause of mesenchymal stem cells death and senescence and a critical issue for survival after transplantation, we propose that melatonin by favoring mitochondria functionality and their transfer through tunneling nanotubes from healthy to suffering cells could improve mesenchymal stem cell-based therapy in a large number of diseases for which basic and clinical trials are underway.

Gastric cancer cell types display distinct proteasome/immunoproteasome patterns associated with migration and resistance to proteasome inhibitors.

PURPOSE: Gastric cancers (GC) display histological and molecular differences. This heterogeneity has limited the development of new therapeutic strategies which requires the identification of the molecular players involved in GC pathogenesis and the investigation of their responsiveness to drugs. Several proteasome subunits have been identified as prognostic markers in GC and their role studied by gene knockdown. However, proteasomes are multi-subunit protein complexes co-existing in multiple forms with distinct activity/specificity and ability to change in response to inhibitors. Information on the role of different proteasome particles in cancer and their relevance as therapeutic targets is limited. METHODS: Based on this evidence, subunit assembly into proteasome complexes and activity were investigated by native PAGE followed by immunoblotting, and by using fluorogenic substrates, respectively. RESULTS: Here we show that GC cell lines with epithelial and/or diffuse Lauren's histotype express different levels of immunoproteasome subunits and equal amounts of constitutive counterparts. Immunoproteasome subunits were highly expressed and preferentially assembled into 19S capped complexes in diffuse-type cells, where most of the activity was catalyzed by the 26S and 30S particles. In epithelial cells, activity appeared equally distributed between 19S- and 11S-capped proteolytic particles. This proteasome pattern was associated with higher resistance of diffuse-type cells to proteasome inhibition. Immunoproteasome inhibition by ONX 0914 did not influence cell viability but affected metastatic cell migration. CONCLUSIONS: These results suggest that pharmacological inhibition of the immunoproteasome may be useful in treating metastatic gastric cancers.

Cytotoxic functions and susceptibility to apoptosis of human CD56(bright) NK cells differentiated in vitro from CD34⁺ hematopoietic progenitors.

Cytotoxic functions and susceptibility to apoptosis are crucial aspects of NK cells suitable to counter cancer after infusion in oncologic patients. To test the feasibility and the usefulness of infusing in vitro generated NK cells, these two features were investigated in NK cells developed in vitro from CD34⁺ hematopoietic progenitors. Purified CD34⁺ cells were cultured for 15-30 days with FLT-3 ligand (FLT3-L) and IL-15 with or without IL-21. To induce terminal differentiation, NK cells were cultured for further 15 days with IL-15, IL-21, or their combination. A CD56(dim) /CD16⁺ NK subset, expressing high level of perforin, granzymes, and LFA-1, appeared early in cultures with FLT3-L, IL-15, and IL-21, but it quickly died, indicating its predisposition to apoptosis. On the contrary, CD56(bright) NK cells generated after 30 days of culture with FLT3-L plus IL-15 did not show a considerable apoptosis, nevertheless only a subset of these cells expressed granzyme-B, perforin, LFA-1, and CD94-CD159a heterodimer, indicating a functional immaturity. Interestingly, further 15 days of culture with IL-21 plus IL-15 did not induce the generation of CD56(dim) cells from the CD56(bright) subset and actually inhibited IL-15-induced maturation/activation of this latter subset. In fact, IL-15 alone upregulated granzyme-B, TRAIL, Fas ligand, CD94-CD159a, LFA-1, CD16, KIRs, and TRAIL-R2 on CD56(bright) NK cells. Our results suggest that during differentiation CD56(bright) NK cells, similarly to mature activated NK cells, become highly cytotoxic and are relatively resistant to apoptosis induced by TNF family members.

The effects of Acyclovir administration to NCI-H1975 non-small cell lung cancer cells.

The biochemical mechanisms by which the antiviral drug Acyclovir (ACV) may induce anticancer effects even without detecting human herpesviruses (HHVs) are still poorly understood. Herein, we investigated for the first time how NCI-H1975 non-small cell lung cancer cells responded in vitro to ACV administration by exploring mitochondrial damage and apoptosis induction. We confirmed ACV ability to cause the inhibition of cancer cell growth even without detecting intracellular HHVs; the drug also significantly inhibited the colony formation capacity of NCI-H1975 cells. Cell cycle analysis revealed an increase of the sub-G1 hypodiploid peak after ACV treatment; the activation of caspase-3 and the presence of DNA laddering sustained the capacity of the drug to induce apoptotic cell death. Regarding mitochondrial toxicity, a reduction of mitochondrial membrane potential, altered mitochondrial size and shape, and mtDNA damage were found after ACV administration. Furthermore, an increment of intracellular reactive oxygen species levels as well as the upregulation of NudT3 involved in DNA repair mechanisms were observed. Altogether, these findings suggest that mitochondria may be possible initial targets and/or sites of ACV cytotoxicity within cancer cells in the absence of intracellular HHVs.

Melatonin Attenuates Ischemic-like Cell Injury by Promoting Autophagosome Maturation via the Sirt1/FoxO1/Rab7 Axis in Hippocampal HT22 Cells and in Organotypic Cultures.

Dysfunctional autophagy is linked to neuronal damage in ischemia/reperfusion injury. The Ras-related protein 7 (Rab7), a member of the Rab family of small GTPases, appears crucial for the progression of the autophagic flux, and its activity is strictly interconnected with the histone deacetylase Silent information regulator 1 (Sirt1) and transcription factor Forkhead box class O1 (FoxO1). The present study assessed the neuroprotective role of melatonin in the modulation of the Sirt1/FoxO1/Rab7 axis in HT22 cells and organotypic hippocampal cultures exposed to oxygen-glucose deprivation followed by reoxygenation (OGD/R). The results showed that melatonin re-established physiological levels of autophagy and reduced propidium iodide-positive cells, speeding up autophagosome (AP) maturation and increasing lysosomal activity. Our study revealed that melatonin modulates autophagic pathways, increasing the expression of both Rab7 and FoxO1 and restoring the Sirt1 expression affected by OGD/R. In addition, the Sirt1 inhibitor EX-527 significantly reduced Rab7, Sirt1, and FoxO1 expression, as well as autolysosomes formation, and blocked the neuroprotective effect of melatonin. Overall, our findings provide, for the first time, new insights into the neuroprotective role of melatonin against ischemic injury through the activation of the Sirt1/FoxO1/Rab7 axis.

3ß-Hydroxy-5ß-hydroxy-B-norcholestane-6ß-carboxaldehyde (SEC-B) Induces Proinflammatory Activation of Human Endothelial Cells Associated with Nitric Oxide Production and Endothelial Nitric Oxide Synthase/Caveolin-1 Dysregulation.

Oxysterols are a family of 27-carbon cholesterol oxidation derivatives found in low-density lipoproteins (LDLs) and atherosclerotic plaques where they trigger several biological responses involved in the initiation and progression of atherosclerosis. Several pieces of evidence suggest that oxysterols contribute to endothelial dysfunction (ED) due to their ability to alter membrane fluidity and cell permeability leading to inflammation, oxidative stress and apoptosis. The present study aimed to investigate the molecular events occurring in human microvascular endothelial cells (HMEC-1) in response to autoxidation-generated 3ß-hydroxy-5ß-hydroxy-B-norcholestane-6ß-carboxaldehyde (SEC-B) exposure. Our results highlight that SEC-B rapidly activates HMEC-1 by inducing oxidative stress, nitric oxide (NO) production and pro-inflammatory cytokine release. Exposure to SEC-B up to 24 h results in persistent accumulation of the vasodilator NO paralleled by an upregulation of the endothelial nitric oxide synthase (eNOS) enzyme and downregulation of Caveolin-1 (Cav-1) protein levels. Moreover, reduced expression and extracellular release of the vasoconstrictor factor endothelin-1 (ET-1) are observed. Furthermore, SEC-B stimulates the expression of the cytokines interleukin-6 (IL-6) and tumor necrosis factor-like weak inducer of apoptosis (TWEAK). This proinflammatory state leads to increased monocyte recruitment on activated HMEC-1 cells. Our findings add new knowledge on the role of SEC-B in ED and further support its potential implication in atherosclerosis.

Melatonin signaling and cell protection function.

Besides its well-known regulatory role on circadian rhythm, the pineal gland hormone melatonin has other biological functions and a distinct metabolism in various cell types and peripheral tissues. In different tissues and organs, melatonin has been described to act as a paracrine and also as an intracrine and autocrine agent with overall homeostatic functions and pleiotropic effects that include cell protection and prosurvival factor. These latter effects, documented in a number of in vitro and in vivo studies, are sustained through both receptor-dependent and -independent mechanisms that control detoxification and stress response genes, thus conferring protection against a number of xenobiotics and endobiotics produced by acute and chronic noxious stimuli. Redox-sensitive components are included in the cell protection signaling of melatonin and in the resulting transcriptional response that involves the control of NF-κB, AP-1, and Nrf2. By these pathways, melatonin stimulates the expression of antioxidant and detoxification genes, acting in turn as a glutathione system enhancer. A further and converging mechanism of cell protection by this indoleamine described in different models seems to lie in the control of damage and signaling function of mitochondria that involves decreased production of reactive oxygen species and activation of the antiapoptotic and redox-sensitive element Bcl2. Recent evidence suggests that upstream components in this mitochondrial route include the calmodulin pathway with its central role in melatonin signaling and the survival-promoting component of MAPKs, ERK1/2. In this review article, we will discuss these and other molecular aspects of melatonin signaling relevant to cell protection and survival mechanisms.

LDL receptors, caveolae and cholesterol in endothelial dysfunction: oxLDLs accomplices or victims?

Oxidized LDLs (oxLDLs) and oxysterols play a key role in endothelial dysfunction and the development of atherosclerosis. The loss of vascular endothelium function negatively impacts vasomotion, cell growth, adhesiveness and barrier functions. While for some of these disturbances, a reasonable explanation can be provided from a mechanistic standpoint, for many others, the molecular mediators that are involved are unknown. Caveolae, specific plasma membrane domains, have recently emerged as targets and mediators of oxLDL-induced endothelial dysfunction. Caveolae and their associated protein caveolin-1 (Cav-1) are involved in oxLDLs/LDLs transcytosis, mainly through the scavenger receptor class B type 1 (SR-B1 or SCARB1). In contrast, oxLDLs endocytosis is mediated by the lectin-like oxidized LDL receptor 1 (LOX-1), whose activity depends on an intact caveolae system. In addition, LOX-1 regulates the expression of Cav-1 and vice versa. On the other hand, oxLDLs may affect cholesterol plasma membrane content/distribution thus influencing caveolae architecture, Cav-1 localization and the associated signalling. Overall, the evidence indicate that caveolae have both active and passive roles in oxLDL-induced endothelial cell dysfunction. First, as mediators of lipid uptake and transfer in the subendothelial space and, later, as targets of changes in composition/dynamics of plasma membrane lipids resulting from increased levels of circulating oxLDLs. Gaining a better understanding of how oxLDLs interact with endothelial cells and modulate caveolae-mediated signalling pathways, leading to endothelial dysfunction, is crucial to find new targets for intervention to tackle atherosclerosis and the related clinical entities. LINKED ARTICLES: This article is part of a themed issue on Oxysterols, Lifelong Health and Therapeutics. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v178.16/issuetoc.

I-152, a supplier of N-acetyl-cysteine and cysteamine, inhibits immunoglobulin secretion and plasma cell maturation in LP-BM5 murine leukemia retrovirus-infected mice by affecting the unfolded protein response.

Excessive production of immunoglobulins (Ig) causes endoplasmic reticulum (ER) stress and triggers the unfolded protein response (UPR). Hypergammaglobulinemia and lymphadenopathy are hallmarks of murine AIDS that develops in mice infected with the LP-BM5 murine leukemia retrovirus complex. In these mice, Th2 polarization and aberrant humoral response have been previously correlated to altered intracellular redox homeostasis. Our goal was to understand the role of the cell's redox state in Ig secretion and plasma cell (PC) maturation. To this aim, LP-BM5-infected mice were treated with I-152, an N-acetyl-cysteine and cysteamine supplier. Intraperitoneal I-152 administration (30 µmol/mouse three times a week for 9 weeks) decreased plasma IgG and increased IgG/Syndecan 1 ratio in the lymph nodes where IgG were in part accumulated within the ER. PC containing cytoplasmic inclusions filled with IgG were present in all animals, with fewer mature PC in those treated with I-152. Infection induced up-regulation of signaling molecules involved in the UPR, i.e. CHAC1, BiP, sXBP-1 and PDI, that were generally unaffected by I-152 treatment except for PDI and sXBP-1, which have a key role in protein folding and PC maturation, respectively. Our data suggest that one of the mechanisms through which I-152 can limit hypergammaglobulinemia in LP-BM5-infected mice is by influencing IgG folding/assembly as well as secretion and affecting PC maturation.