Cells In Specific Area Research Articles

Functionalization of TiO2 sol-gel derived films for cell confinement

The neuroscience field has increased enormously over the last decades, achieving the possible real application of neuronal cultures not only for reproducing neural architectures resembling in vivo tissues, but also for the development of functional devices. In this context, surface patterning for cell confinement is crucial, and new active materials together with new protocols for preparing substrates suitable for confining cells, guiding their processes in the desired configuration are extremely appreciated. Here, TiO2 sol-gel derived films were selected as proof-of-concept materials to grow neurons in suitable confined configurations, taking advantage of the biocompatible properties of modified TiO2 substrates. TiO2 sol-gel derived films were made compatible with the growth of neurons thanks to a stable and controlled poly-lysine coating, obtained by silanization chemistry and streptavidin-biotin interactions. Moreover, a spotting protocol, here described and optimized, allowed the simple preparation of arrays of neurons, where cell adhesion was guided in specific areas and the neurites development driven in the desired arrangement. The resulting arrays were successfully tested for the growth and differentiation of neurons, demonstrating the possible adhesion of cells in specific areas of the film, therefore paving the way to applications such as the direct growth of excitable cells nearby electrodes of devices, with an evident enhancement of cell-electrodes communication.

A State of the Art of Antioxidant Properties of Curcuminoids in Neurodegenerative Diseases.

Neurodegenerative diseases represent a set of pathologies characterized by an irreversible and progressive, and a loss of neuronal cells in specific areas of the brain. Oxidative phosphorylation is a source of energy production by which many cells, such as the neuronal cells, meet their energy needs. Dysregulations of oxidative phosphorylation induce oxidative stress, which plays a key role in the onset of neurodegenerative diseases such as Alzheimer’s disease (AD), Parkinson’s disease (PD), and amyotrophic lateral sclerosis (ALS). To date, for most neurodegenerative diseases, there are no resolute treatments, but only interventions capable of alleviating the symptoms or slowing the course of the disease. Therefore, effective neuroprotection strategies are needed. In recent years, natural products, such as curcuminoids, have been intensively explored and studied for their therapeutic potentials in several neurodegenerative diseases. Curcuminoids are, nutraceutical compouns, that owen several therapeutic properties such as anti-oxidant, anti-inflammatory and neuroprotective effects. In this context, the aim of this review was to provide an overview of preclinical and clinical evidence aimed to illustrate the antioxidant effects of curcuminoids in neurodegenerative diseases. Promising results from preclinical studies encourage the use of curcuminoids for neurodegeneration prevention and treatment.

GABAARα2 is Decreased in the Axon Initial Segment of Pyramidal Cells in Specific Areas of the Prefrontal Cortex in Autism

Some forms of Autism Spectrum Disorder, a neurodevelopmental syndrome characterized by impaired communication and social skills as well as repetitive behaviors, are purportedly associated with dysregulation of the excitation/inhibition balance in the cerebral cortex. Through human postmortem tissue analysis, we previously found a significant decrease in the number of a gamma-aminobutyric acid (GABA)ergic interneuron subtype, the chandelier (Ch) cell, in the prefrontal cortex of subjects with autism. Ch cells exclusively target the axon initial segment (AIS) of excitatory pyramidal (Pyr) neurons, and a single Ch cell forms synapses on hundreds of Pyr cells, indicating a possible role in maintaining electrical balance. Thus, we herein investigated this crucial link between Ch and Pyr cells in the anatomy of autism neuropathology by examining GABA receptor protein expression in the Pyr cell AIS in subjects with autism. We collected tissue from the prefrontal cortex (Brodmann Areas (BA) 9, 46, and 47) of 20 subjects with autism and 20 age- and sex-matched control subjects. Immunohistochemical staining with antibodies against the GABAA receptor subunit α2 (GABAARα2) – the subunit most prevalent in the Pyr cell AIS – revealed a significantly decreased GABAARα2 protein in the Pyr cell AIS in supragranular layers of prefrontal cortical areas BA9 and BA47 in autism. Downregulated GABAARα2 protein in the Pyr cell AIS may result from decreased GABA synthesis in the prefrontal cortex of subjects with autism, and thereby contribute to an excitation/inhibition imbalance. Our findings support the potential for GABA receptor agonists asa therapeutic tool for autism.

Rationally Designed Peptide Interface for Potential Modulated Cell Adhesion and Migration

Conformational changes of peptides are critically important in the control of their biological activities. Here, a quaternary ammonium group‐terminated RGD‐containing peptide (RGD‐NMe3) is designed, which may undergo reversible conformational switch upon different electrochemical potentials. Potential responsive peptide interfaces are constructed on gold substrates with RGD‐NMe3 in a tetra (ethylene glycol) background. It is demonstrated that by applying positive and negative potentials, the RGD peptide can be reversibly switched between linear and cyclic conformation, which can be used in reversible controlling of cell adhesion/migration on the interface. Furthermore, by combining microfluidics, adhesion of the cells in specific areas on the surface and subsequent directional migration of the cells can be controlled. It is believed that this straightforward potential modulation mechanism for peptide conformation control may find a wide use in design responsive peptide interfaces.

Use of <em>In vivo</em> Imaging to Monitor the Progression of Experimental Mouse Cytomegalovirus Infection in Neonates

Human Cytomegalovirus (HCMV or HHV-5) is a life-threatening pathogen in immune-compromised individuals. Upon congenital or neonatal infection, the virus can infect and replicate in the developing brain, which may induce severe neurological damage, including deafness and mental retardation. Despite the potential severity of the symptoms, the therapeutic options are limited by the unavailability of a vaccine and the absence of a specific antiviral therapy. Furthermore, a precise description of the molecular events occurring during infection of the central nervous system (CNS) is still lacking since observations mostly derive from the autopsy of infected children. Several animal models, such as rhesus macaque CMV, have been developed and provided important insights into CMV pathogenesis in the CNS. However, despite its evolutionary proximity with humans, this model was limited by the intracranial inoculation procedure used to infect the animals and consistently induce CNS infection. Furthermore, ethical considerations have promoted the development of alternative models, among which neonatal infection of newborn mice with mouse cytomegalovirus (MCMV) has recently led to significant advances. For instance, it was reported that intraperitoneal injection of MCMV to Balb/c neonates leads to infection of neurons and glial cells in specific areas of the brain. These findings suggested that experimental inoculation of mice might recapitulate the deficits induced by HCMV infection in children. Nevertheless, a dynamic analysis of MCMV infection of neonates is difficult to perform because classical methodology requires the sacrifice of a significant number of animals at different time points to analyze the viral burden and/or immune-related parameters. To circumvent this bottleneck and to enable future investigations of rare mutant animals, we applied in vivo imaging technology to perform a time-course analysis of the viral dissemination in the brain upon peripheral injection of a recombinant MCMV expressing luciferase to C57Bl/6 neonates.

Use of <em>In vivo</em> Imaging to Monitor the Progression of Experimental Mouse Cytomegalovirus Infection in Neonates

Human Cytomegalovirus (HCMV or HHV-5) is a life-threatening pathogen in immune-compromised individuals. Upon congenital or neonatal infection, the virus can infect and replicate in the developing brain, which may induce severe neurological damage, including deafness and mental retardation. Despite the potential severity of the symptoms, the therapeutic options are limited by the unavailability of a vaccine and the absence of a specific antiviral therapy. Furthermore, a precise description of the molecular events occurring during infection of the central nervous system (CNS) is still lacking since observations mostly derive from the autopsy of infected children. Several animal models, such as rhesus macaque CMV, have been developed and provided important insights into CMV pathogenesis in the CNS. However, despite its evolutionary proximity with humans, this model was limited by the intracranial inoculation procedure used to infect the animals and consistently induce CNS infection. Furthermore, ethical considerations have promoted the development of alternative models, among which neonatal infection of newborn mice with mouse cytomegalovirus (MCMV) has recently led to significant advances. For instance, it was reported that intraperitoneal injection of MCMV to Balb/c neonates leads to infection of neurons and glial cells in specific areas of the brain. These findings suggested that experimental inoculation of mice might recapitulate the deficits induced by HCMV infection in children. Nevertheless, a dynamic analysis of MCMV infection of neonates is difficult to perform because classical methodology requires the sacrifice of a significant number of animals at different time points to analyze the viral burden and/or immune-related parameters. To circumvent this bottleneck and to enable future investigations of rare mutant animals, we applied in vivo imaging technology to perform a time-course analysis of the viral dissemination in the brain upon peripheral injection of a recombinant MCMV expressing luciferase to C57Bl/6 neonates.

Region Specific Hypothalamic Neuronal Activation and Endothelial Cell Proliferation in Response to Electroconvulsive Seizures

Major depression is often associated with disturbances in basal biological functions regulated by the hypothalamus. Electroconvulsive therapy (ECT), an efficient anti-depressant treatment, alters the activity of hypothalamic neurons. We have previously shown an increased proliferation of endothelial cells in specific areas of the rat hippocampus in response to electroconvulsive seizure (ECS) treatment, an animal model for ECT. Here we examine the effect of ECS treatment on neuronal activation and endothelial cell proliferation in mid-hypothalamus. Rats received one daily ECS treatment for 5 days and cell proliferation was detected by bromodeoxyuridine (BrdU). The number of cells double-labeled for BrdU and the endothelial cell marker rat endothelial cell antigen-1 was determined. Neuronal activation in response to acute ECS treatment was detected as c-Fos immunoreactivity in an additional experiment. We demonstrate a correlating pattern of increases in neuronal activation and increased endothelial cell proliferation in the paraventricular nucleus, the supraoptic nucleus, and the ventromedial nucleus of the hypothalamus after ECS treatment. Hypothalamic areas with the largest increase in neuronal activation after ECS treatment exhibit increased endothelial cell proliferation. We suggest that similar angiogenic responses to ECT might counteract hypothalamic dysfunction in depressive disorder.

Localization of renin expressing cells in the brain, by use of a REN-eGFP transgenic model.

Immunoreactive renin has been reported in the hypothalamus and cerebellar cortex in the rodent brain and in neurons in all areas of the human brain. Despite these observations and the clear documentation of the expression of the other renin-angiotensin system genes in the brain, the notion that renin is endogenously expressed in the brain remains very controversial and undefined. This controversy no doubt arises because the level of renin expression in the brain is below the detection threshold of most standard assays. A transgenic mouse expressing enhanced green fluorescence protein (eGFP) under the control of the mouse renin promoter was recently reported. This model expresses eGFP in the kidney, which responds appropriately to both developmental and physiological stimuli. We therefore used eGFP as a sensitive marker to identify renin-expressing cells in the brain. We identified eGFP-containing cells in specific areas of the brain, including cerebellum, hippocampus, dorsal motor nucleus of the vagus, inferior olivary nucleus, reticular formation, rostral ventrolateral medulla, central nucleus of the amygdala, lateral parabrachial nucleus, mesencephalic trigeminal nucleus, bed nucleus of stria terminalis, and subfornical organ. By colabeling with neuron- or glia (astrocytes or oligodendrocytes)-specific antisera, we have determined the eGFP-positive cells to be mainly neuronal. These findings therefore strongly support the primary expression of renin mRNA in the brain in regions controlling cardiovascular function.

Adolescent development alters stressor-induced Fos immunoreactivity in rat brain

Adult-typical behavioural responses to environmental challenges as well as the stressor responsiveness of several neural systems emerge over adolescent development. The present study was undertaken to determine whether stressors might activate different neural populations in adult vs juvenile male rats. Fos-immunoreactivity was determined in various forebrain nuclei following 15 min or 2 h of restraint in 28- and 60-day-old male rats (representing late juvenile and young adult ages, respectively) and compared to non-restrained control animals at each age. Few Fos-positive cells were identifed in unrestrained controls at either age. Restraint, however, induced the production of Fos in several areas. Fos immunoreactivity was marked in parvocellular regions of the paraventricular nucleus of the hypothalamus following both restraint periods and at both ages, an observation consistent with previous observations that restraint increases plasma corticosterone at both ages. And at both ages, Fos immunoreactivity was evident in magnocellular regions of the hypothalamus only following the longer restraint period. Fos immunoreactivity, however, clearly varied as a function of adolescent age in several regions. Moderate to intense Fos immunoreactivity was observed in adults in all divisions of the anterior olfactory nucleus, cortical and medial amygdaloid nuclei, pyriform cortex and tenia tecta. In contrast to the adult, only a few Fos positive cells were observed in any of these regions in juveniles. Exposure to the same stressor induced Fos in a broader spectrum of neurons in young adult than in juvenile male rats. The lack of Fos-positive cells in specific areas of juveniles may relate to maturation in specific amygdaloid nuclei, which project to many of the other regions that showed age-related differences in Fos production. The emergence over adolescence of Fos-positive cells in specific areas in response to stressors may underlie the emergence of adult-typical behavioural and neural stressor-responsiveness.

Melatonin: Its intracellular and genomic actions

Besides its actions via membrane receptors on cells in specific areas of the brain and peripheral tissues, melatonin also has direct intracellular actions. The neurohormone melatonin is a potent free radical scavenger in vitro and a powerful antioxidant in vivo. Studies to date indicate that it is a better scavenger of the highly toxic hydroxyl and peroxyl radicals than some other known compounds. Melatonin also prevents the toxicity of singlet oxygen and stimulates the antioxidative enzyme, glutathione peroxidase. Considering its varied and potent antioxidant capability, it is possible that melatonin is an essential element of the antioxidant defense system of organisms. Besides these direct intracellular actions of melatonin, it was recently discovered that the indole also has binding sites in the nucleus of many cells. Melatonin's genomic actions are believed to follow its binding to these nuclear loci.