Stefano Loizzo

Potential Therapeutic Effects of Vitamin E and C on Placental Oxidative Stress Induced by Nicotine: An In Vitro Evidence

There have been a few studies that examined the oxidative stress effects of nicotine during pregnancy and lactation. The adverse effect of prenatal smoking exposure on human fetal development and growth has been a major public health issue. Active or passive smoking during pregnancy can result in a wide variety of adverse outcomes, including intrauterine growth retardation (IUGR), prematurity, stillbirth, and the sudden infant death syndrome. Smoking in pregnancy has also been associated with an increased risk of attention deficit and learning problems in childhood. Some studies argued that as a principal component of tobacco smoke, nicotine alone is responsible for the majority of negative reproductive outcomes. Nicotine and its major metabolite cotinine can cross the placental barrier. The level of nicotine in fetal tissues was found to be equal to or greater than the plasma nicotine level in the mothers. The oxidative stress induce by nicotine has been increasingly postulated as a major contributor to endothelial dysfunction. A large body of research has investigated the potential role of antioxidant nutrients in the prevention of endothelial dysfunction in women. Therefore, the present study was undertaken to assess the potential benefit of antioxidant supplementation on markers of placental oxidative stress in an in vitro model of endothelial dysfunction induced by nicotine, since it was previously found that nicotine is able to trigger the placental secretion of stress molecules. In this regard, we evaluated the effects of vitamin C, vitamin E and N-acetylcysteine (NAC), alone or in combination, in placental villi culture after exposure to nicotine. The effect of antioxidant nutrients on trophoblast cells proliferation and vitality was also evaluated. The results obtained suggest that in a patho-physiological condition, such as endothelial dysfunction induced by nicotine, the deleterious effect of reactive oxygen species may be counteracted by an antioxidant therapy, and there is the need to investigate the optimum dosing and timing of antioxidants administration, since an inappropriate antioxidant treatment in pregnant women may have deleterious consequences, reducing placental cells proliferation until to cell death.

Naloxone prevents cell-mediated immune alterations in adult mice following repeated mild stress in the neonatal period

Mild stress plus mild pain (solvent injection) applied daily to neonatal mice induces hormonal, behavioural and metabolic changes perduring in the adult life. We investigated whether daily mild stress to neonatal mice induces also long‐term defined changes of immune response, and whether immune changes are prevented through repeated administration of the opioid antagonist naloxone. Mild stress plus solvent injection administered from birth to the 21st postnatal day causes not only behavioural and metabolic changes, but also long‐term (up to 110 days of life) splenocytes modifications, consisting in: increased release of the Th‐1 type cytokines interleukin‐2 (IL‐2) (from an average of 346 to 788 pg ml−1), interferon‐γ (from 1770 to 3942) and tumour necrosis factor‐α (from 760 to 1241); decreased release of the Th‐2 type cytokines IL‐4 (from 49.1 to 28.4) and IL‐10 (from 1508 to 877). Moreover, enhanced natural killer‐cell activity; enhanced proliferative splenocytes properties in resting conditions and following phytohemoagglutinin and concanavalin‐A stimulation are observed. Immunological, behavioural and metabolic changes are prevented by the opioid antagonist (−)naloxone (1 mg kg−1 per day s.c., administered instead of solvent) but not by the biologically inactive enantiomorph (+)naloxone. In conclusion, endogenous opioid systems sensitive to naloxone are involved in long‐lasting enhancement of the Th‐1 type cytokines and cell‐mediated immunological response caused by repeated mild stress administered postnatally.

Overweight and Metabolic and Hormonal Parameter Disruption Are Induced in Adult Male Mice by Manipulations During Lactation Period

Neonatal manipulations (10 min of maternal separation plus s.c. sham injection, daily for the first 21 d of life) determine overweight in male adult mice. In this work, we investigated the mechanisms underlying mild obesity and the alteration of caloric balance. Neonatally manipulated mice become overweight after onset of maturity, showing increased fat tissue and hypertrophic epididymal adipocytes. Increase in body weight occurs in the presence of a small increase in daily food intake (significant only in the adult period) and the absence of a decrease in spontaneous locomotor activity, while the calculated caloric efficiency is higher in manipulated mice, especially in adulthood. Fasting adult animals show hyperglycemia, hyperinsulinemia, hypertriglyceridemia, hypercholesterolemia, and hyperleptinemia. Soon after weaning and in the adulthood, plasma corticosterone and adrenocorticotropin (ACTH) are also significantly increased. Thus, neonatal manipulations in nongenetically susceptible male mice program mild obesity, with metabolic and hormonal alterations that are similar to those found in experimental models of diabetes mellitus, suggesting that this metabolic derangement may have at least part of its roots early on in life and, more interestingly, that psychological and nociceptive stimuli induce these features.

Abnormal NMDA receptor function exacerbates experimental autoimmune encephalomyelitis

Glutamate transmission is dysregulated in both multiple sclerosis (MS) and experimental autoimmune encephalomyelitis (EAE), the animal model of MS. A characteristic of EAE is increased glutamate transmission associated with up‐regulation of AMPA receptors. However, little is known about the role of NMDA receptors in the synaptic modifications induced by EAE.

CNF1 increases brain energy level, counteracts neuroinflammatory markers and rescues cognitive deficits in a murine model of Alzheimer's disease.

Overexpression of pro-inflammatory cytokines and cellular energy failure are associated with neuroinflammatory disorders, such as Alzheimers disease. Transgenic mice homozygous for human ApoE4 gene, a well known AD and atherosclerosis animal model, show decreased levels of ATP, increased inflammatory cytokines level and accumulation of beta amyloid in the brain. All these findings are considered responsible for triggering cognitive decline. We have demonstrated that a single administration of the bacterial E. coli protein toxin CNF1 to aged apoE4 mice, beside inducing a strong amelioration of both spatial and emotional memory deficits, favored the cell energy restore through an increment of ATP content. This was accompanied by a modulation of cerebral Rho and Rac1 activity. Furthermore, CNF1 decreased the levels of beta amyloid accumulation and interleukin-1β expression in the hippocampus. Altogether, these data suggest that the pharmacological modulation of Rho GTPases by CNF1 can improve memory performances in an animal model of Alzheimers disease via a control of neuroinflammation and a rescue of systemic energy homeostasis.

A chronic implant to record electroretinogram, visual evoked potentials and oscillatory potentials in awake, freely moving rats for pharmacological studies.

Electroretinogram (ERG), widely used to study the pharmacological effects of drugs in animal models (e.g., diabetic retinopathy), is usually recorded in anesthetized rats. We report here a novel simple method to obtain chronic implantation of electrodes for simultaneous recording at the retinal and cortical levels in freely moving, unanesthetized animals. We recorded cortical (VEPs) and retinal (ERGs) responses evoked by light (flash) stimuli in awake rats and compared the results in the same rats anesthetized with urethane (0.6 mg/kg) before and after the monocular administration of scopolamine methyl bromide (1‰solution). We also compared the retinal responses with those derived from a classic acute corneal electrode. Anesthesia induced consistent changes of several VEP and ERG parameters like an increase of both latency and amplitude. In particular, the analysis of the variation of latency, amplitude, and spectral content of rapid oscillatory potentials could be important for a functional evaluation of the visual system in unanesthetized versus anesthetized animals.

The Cytotoxic Necrotizing Factor 1 from E. Coli: A Janus Toxin Playing with Cancer Regulators

Certain strains of Escherichia coli have been indicated as a risk factor for colon cancer. E. coli is a normal inhabitant of the human intestine that becomes pathogenic, especially in extraintestinal sites, following the acquisition of virulence factors, including the protein toxin CNF1. This Rho GTPases-activating toxin induces dysfunctions in transformed epithelial cells, such as apoptosis counteraction, pro-inflammatory cytokines’ release, COX2 expression, NF-kB activation and boosted cellular motility. As cancer may arise when the same regulatory pathways are affected, it is conceivable to hypothesize that CNF1-producing E. coli infections can contribute to cancer development. This review focuses on those aspects of CNF1 related to transformation, with the aim of contributing to the identification of a new possible carcinogenic agent from the microbial world.

Dynamic NAD(P)H post-synaptic autofluorescence signals for the assessment of mitochondrial function in a neurodegenerative disease: Monitoring the primary motor cortex of G93A mice, an amyotrophic lateral sclerosis model

Abnormal mitochondrial function was reported in patients and models for amyotrophic lateral sclerosis (ALS). It is therefore important to set up sensitive tools for the monitoring of active agents that enhance energy metabolism delay onset, and extend lifespan of transgenic G93A-SOD1 ALS mice. In this report, primary motor cortex slices from G93A mice at different stages of disease were studied, using NAD(P)H autofluorescence post-synaptic signals following ultraviolet stimuli, as a probe to evaluate mitochondrial function. We observed consistent age-related alterations of responses in G93A primary motor cortex slices versus controls. We conclude that NAD(P)H autofluorescence post-synaptic signal is a highly sensitive real-time technique to detect mitochondrial function failure in primary cortex from living tissues.

Modulation of flash stimulation intensity and frequency: effects on visual evoked potentials and oscillatory potentials recorded in awake, freely moving mice.

Visual evoked potentials (VEP) responses to flash stimulation at nine intensities, from 0.611 to 945.6 cd/m(2)*s, and two frequencies (0.2 and 1 Hz) were recorded and oscillatory potentials (OPs) extracted after digital 50-Hz high pass filtering in unanaesthetized unrestrained mice. Both VEP and OPs morphology were replicable for all conditions and were similar to values reported in the literature. In particular OPs spectral analysis showed that the main frequency component remained stable at 66-77 Hz, for both stimulation frequencies, although it displayed an increase in amplitude, as a function of stimulus intensity. OPs amplitude at 1 Hz versus 0.2 Hz stimulus frequency was higher after taking into account the different noise contributions in the two conditions. Root mean square values calculated at selected time windows, revealed that, at 1 Hz, the main contribution to OPs occurs at the onset of the response (14-27 ms) while, at 0.2 Hz, the higher RMS was recorded later (42-56 ms). This difference accounts for the longer duration of the oscillatory event in the 0.2-Hz condition and suggests that oscillatory activity, modulated and carried along the visual pathway, is recorded at the cortical electrode after further elaboration at the cortical/subcortical level, depending on stimulus properties.

Mode of action of fibrous amphiboles: the case of Biancavilla (Sicily, Italy)

BACKGROUND The inhalation of fibrous amphiboles can result in pulmonary fibrosis, lung cancer and mesothelioma. Although these fibres have the same disease-causing potential, their different morphologies and chemical composition can determine different biological activities. An unusual cluster of mesothelioma was evidenced in Biancavilla (Sicily) where no inhabitant had been significantly exposed to asbestos. OBJECTIVE We herein discuss the mechanism of action of amphiboles, focusing on the fibres identified in the study area. RESULTS Human lung carcinoma cells have been exposed to two different materials: prismatic fluoro-edenite and fibres with fluoro-edenitic composition. Only in the second case, they exhibit features typical of transformed cells, such as multinucleation, prosurvival activity and pro-inflammatory cytokine release. Accordingly, in vivo studies demonstrated that the fibrous sample only could induce a mesotheliomatogenic effect. CONCLUSIONS Fibres with fluoro-edenitic composition behave similarly to the asbestos crocidolite, whose connection with inflammation and lung cancer is well established.