Flavopiridol: An Old Drug With New Perspectives? Implication for Development of New Drugs.

Glioblastoma, the most common brain tumor, is characterized by high proliferation rate, invasion, angiogenesis, and chemo- and radio-resistance. One of most remarkable feature of glioblastoma is the switch toward a glycolytic energetic metabolism that leads to high glucose uptake and consumption and a strong production of lactate. Activation of several oncogene pathways like Akt, c-myc, and ras induces glycolysis and angiogenesis and acts to assure glycolysis prosecution, tumor proliferation, and resistance to therapy. Therefore, the high glycolytic flux depends on the overexpression of glycolysis-related genes resulting in an overproduction of pyruvate and lactate. Metabolism of glioblastoma thus represents a key issue for cancer research. Flavopiridol is a synthetic flavonoid that inhibits a wide range of Cyclin-dependent kinase, that has been demonstrate to inactivate glycogen phosphorylase, decreasing glucose availability for glycolysis. In this work the study of glucose metabolism upon flavopiridol treatment in the two different glioblastoma cell lines. The results obtained point towards an effect of flavopiridol in glycolytic cells, thus suggesting a possible new use of this compound or flavopiridol-derived formulations in combination with anti-proliferative agents in glioblastoma patients. J. Cell. Physiol. 232: 312-322, 2017. © 2016 Wiley Periodicals, Inc.

The PPARß/δ Agonist GW0742 Induces Early Neuronal Maturation of Cortical Post-Mitotic Neurons: Role of PPARß/δ in Neuronal Maturation.

Increasing evidences support that signaling lipids participate in synaptic plasticity and cell survival, and that the lipid signaling is closely associated with neuronal differentiation, learning, and memory and with pathologic events, such as epilepsy and Alzheimer's disease. The Peroxisome Proliferator-Activated Receptors (PPAR) are strongly involved in the fatty acid cell signaling, as many of the natural lypophylic compounds are PPAR ligands. We have previously shown that PPARß/δ is the main isotype present in cortical neuron primary cultures and that during neuronal maturation, PPARß/δ is gradually increased and activated. To get more insight into the molecular mechanism by which PPARß/δ may be involved in neuronal maturation processes, in this work a specific PPARß/δ agonist, GW0742 was used administered alone or in association with a specific PPARß/δ antagonist, the GSK0660, and the parameters involved in neuronal differentiation and maturation were assayed. The data obtained demonstrated the strong involvement of PPARß/δ in neuronal maturation, triggering the agonist an anticipation of neuronal differentiation, and the antagonist abolishing the observed effects. These effects appear to be mediated by the activation of BDNF pathway.

Cocoa powder triggers neuroprotective and preventive effects in a human Alzheimer's disease model by modulating BDNF signaling pathway.

The molecular mechanisms linking Aß to the onset of neurotoxicity are still largely unknown, but several lines of evidence point to reactive oxygen species, which are produced even under the effect of nanomolar concentrations of soluble Aß-oligomers. The consequent oxidative stress is considered as the mediator of a cascade of degenerative events in many neurological disorders. Epidemiological studies indicate that dietary habits and antioxidants from diet can influence the incidence of neurodegenerative disorders such as Alzheimer's and Parkinson's diseases. In the recent years, a number of reviews have reported on neuroprotective effects of polyphenols in cell and animal models. However, the majority of these studies have focused only on the anti-oxidant properties of these compounds and less on the mechanism/s of action at cellular level. In this work we investigated the effect of cocoa polyphenolic extract on a human AD in vitro model. The results obtained, other than confirming the anti-oxidant properties of cocoa, demonstrate that cocoa polyphenols triggers neuroprotection by activating BDNF survival pathway, both on Aß plaque treated cells and on Aß oligomers treated cells, resulting in the counteraction of neurite dystrophy. On the light of the results obtained the use of cocoa powder as preventive agent for neurodegeneration is further supported.

Expression of the Nrf2-system at the blood-CSF barrier is modulated by neonatal inflammation and hypoxia-ischemia.

Transcription factor NF-E2-related factor-2 (Nrf2) is a key regulator of endogenous anti-oxidant systems shown to play a neuroprotective role in the adult by preserving blood-brain barrier function. The choroid plexus, site for the blood-CSF barrier, has been suggested to be particularly important in maintaining brain barrier function in development. We investigated the expression of Nrf2- and detoxification-system genes in choroid plexus following systemic LPS injections, unilateral cerebral hypoxia-ischemia (HI) as well as the combination of LPS and HI (LPS/HI). Plexuses were collected at different time points after LPS, HI and LPS/HI in 9-day old mice. mRNA levels of Nrf2 and many of its target genes were analyzed by quantitative PCR. Cell death was analyzed by caspase-3 immunostaining and TUNEL. LPS caused down-regulation of the Nrf2-system genes while HI increased expression at earlier time points. LPS exposure prior to HI prevented many of the HI-induced gene increases. None of the insults resulted in any apparent cell death to choroidal epithelium. These data imply that the function of the inducible anti-oxidant system in the choroid plexus is down-regulated by inflammation, even if choroid cells are not structurally damaged. Further, LPS prevented the endogenous antioxidant response following HI, suggesting the possibility that the choroid plexus may be at risk if LPS is united with an insult that increases oxidative stress such as hypoxia-ischemia.

p73 and p63 regulate the expression of fibroblast growth factor receptor 3.

p53, p63 and p73 make a family of transcription factors that play a vital role in development and cancer. All p53 family members have more than one promoter producing Transactivating (TA) and Dominant Negative (DeltaN) isoforms and their mRNAs are subjected to extensive splicing at 3' end to produce multiple protein products. p53 is usually inactivated by point mutations during tumorigenesis, whereas the expression levels and p63 and p73 are modulated to give tumor cells a selective advantage. In this study, aiming to find novel targets of the p53 family members, we identified FGFR3 as a gene transcriptionally controlled by p63 and p73. FGFR3 has been implicated in development and tumor biology as activating mutations of this gene was described in skeletal disorders, non-invasive skin conditions and superficial bladder cancers. We found that TAp73, TAp63 and DeltaNp63 was capable of inducing FGFR3. siRNA mediated downregulation of DeltaNp63 decreased endogenous FGFR3 protein levels. Our findings of this new link between p53 family proteins and FGFR3 may help understanding the transition of superficial bladder cancers to an invasive phenotype.

Corrigendum to "PPARs in Human Neuroepithelial Tumors: PPAR Ligands as Anticancer Therapies for the Most Common Human Neuroepithelial Tumors".

[This corrects the article DOI: 10.1155/2010/427401.].

Fifty hertz extremely low-frequency electromagnetic field causes changes in redox and differentiative status in neuroblastoma cells.

The current study was designed to establish whether extremely low-frequency electromagnetic fields might affect neuronal homeostasis through redox-sensitive mechanisms. To this end, intracellular reactive oxygen species production, antioxidant and glutathione-based detoxifying capability and genomic integrity after extremely low-frequency electromagnetic fields exposure were investigated. Moreover, we also studied potential extremely low-frequency electromagnetic fields-dependent changes in the proliferative and differentiative cellular status. Results seem to support redox-mediated extremely low-frequency electromagnetic fields effects on biological models as, although no major oxidative damage was detected, after exposure we observed a positive modulation of antioxidant enzymatic expression, as well as a significant increase in reduced glutathione level, indicating a shift of cellular environment towards a more reduced state. In addition, extremely low-frequency electromagnetic fields treatment induced a more differentiated phenotype as well as an increased expression in peroxisome proliferators-activated receptor isotype beta, a class of transcription factors related to neuronal differentiation and cellular stress response. As second point, to deepen how extremely low-frequency electromagnetic fields treatment could affect neuroblastoma cell antioxidant capacity, we examined the extremely low-frequency electromagnetic fields-dependent modifications of cell susceptibility to pro-oxidants. Results clearly showed that 50 Hz extremely low-frequency electromagnetic fields exposure reduces cell tolerance towards oxidative attacks.

LPS-induced inflammatory response triggers cell cycle reactivation in murine neuronal cells through retinoblastoma proteins induction.

Cell cycle reactivation in adult neurons is an early hallmark of neurodegeneration. The lipopolysaccharide (LPS) is a well-known pro-inflammatory factor that provokes neuronal cell death via glial cells activation. The retinoblastoma (RB) family includes RB1/p105, retinoblastoma-like 1 (RBL1/p107), and retinoblastoma-like 2 (Rb2/p130). Several studies have indicated that RB proteins exhibit tumor suppressor activities, and play a central role in cell cycle regulation. In this study, we assessed LPS-mediated inflammatory effect on cell cycle reactivation and apoptosis of neuronally differentiated cells. Also, we investigated whether the LPS-mediated inflammatory response can influence the function and expression of RB proteins. Our results showed that LPS challenges triggered cell cycle reactivation of differentiated neuronal cells, indicated by an accumulation of cells in S and G2/M phase. Furthermore, we found that LPS treatment also induced apoptotic death of neurons. Interestingly, we observed that LPS-mediated inflammatory effect on cell cycle re-entry and apoptosis was concomitant with the aberrant expression of RBL1/p107 and RB1/p105. To the best of our knowledge, our study is the first to indicate a role of LPS in inducing cell cycle re-entry and/or apoptosis of differentiated neuronal cells, perhaps through mechanisms altering the expression of specific members of RB family proteins. This study provides novel information on the biology of post-mitotic neurons and could help in identifying novel therapeutic targets to prevent de novo cell cycle reactivation and/or apoptosis of neurons undergoing neurodegenerative processes.

MicroRNAs: A Puzzling Tool in Cancer Diagnostics and Therapy.

MicroRNAs (miRNAs) constitute a dominating class of small RNAs that regulate diverse cellular functions. Due the pivotal role of miRNAs in biological processes, a deregulated miRNA expression is likely involved in human cancers. MicroRNAs possess tumor suppressor capability, as well as display oncogenic characteristics. Interestingly, miRNAs exist in various biological fluids as circulating entities. Changes in the profile of circulating miRNAs are indicative of pathophysiological conditions in human cancer. This concept has led to consider circulating miRNAs valid biomarkers in cancer diagnostics. Furthermore, current research promotes the use of miRNAs as a target in cancer therapy. However, miRNAs are an evolving research field. Although miRNAs have been demonstrated to be potentially valuable tools both in cancer diagnosis and treatment, a greater effort should be made to improve our understanding of miRNAs biology. This review describes the biology of microRNAs, emphasizing on the use of miRNAs in cancer diagnostics and therapy.

GSK3ß inhibition protects the immature brain from hypoxic-ischaemic insult via reduced STAT3 signalling.

Hypoxic-ischaemic (HI) injury is an important cause of neurological morbidity in neonates. HI leads to pathophysiological responses, including inflammation and oxidative stress that culminate in cell death. Activation of glycogen synthase kinase 3ß (GSK3ß) and the signal transducer and activator of transcription (STAT3) promotes brain inflammation. The purpose of this study was to test whether inhibition of GSK3ß signalling protects against neonatal HI brain injury. Mice were subjected to HI at postnatal day (PND) 9 and treated with a selective GSK3ß inhibitor, SB216763. Brain injury and caspase-3 activation, anti-oxidant and inflammatory mRNA responses and activation of STAT3 were analysed. Our results show that HI reduced phosphorylation of GSK3ß, thus promoting its kinase activity. The GSK3ß inhibitor reduced caspase-3 activation and neuronal cell death elicited by HI and reverted the effects of HI on gene expression of the anti-oxidant enzyme sod2 and mitochondrial factor pgc1α. The HI insult activated STAT3 in glial cells and GSK3ß inhibition attenuated STAT3 phosphorylation and its nuclear translocation following HI. Further, GSK3ß inhibition reduced HI-induced gene expression of pro-inflammatory cytokines tnfα and Il-6, while promoted the anti-inflammatory factor Il-10. In summary, data show that GSK3ß inhibition is neuroprotective in neonatal HI brain injury likely via reduced pro-inflammatory responses by blocking STAT3 signalling. Our study suggests that pharmacological interventions built upon GSK3ß silencing strategies could represent a novel therapy in neonatal brain injury.

Expression of tight junction proteins and transporters for xenobiotic metabolism at the blood-CSF barrier during development in the nonhuman primate (P. hamadryas).

The choroid plexus (CP) is rich in barrier mechanisms including transporters and enzymes which can influence drug disposition between blood and brain. We have limited knowledge of their state in fetus. We have studied barrier mechanisms along with metabolism and transporters influencing xenobiotics, using RNAseq and protein analysis, in the CP during the second-half of gestation in a nonhuman primate (Papio hamadryas). There were no differences in the expression of the tight-junctions at the CP suggesting a well-formed fetal blood-CSF barrier during this period of gestation. Further, the fetal CP express many enzymes for phase I-III metabolisms as well as transporters suggesting that it can greatly influence drug disposition and has a significant machinery to deactivate reactive molecules with only minor gestational changes. In summary, the study suggests that from, at least, midgestation, the CP in the nonhuman primate is restrictive and express most known genes associated with barrier function and transport.

Brain barrier properties and cerebral blood flow in neonatal mice exposed to cerebral hypoxia-ischemia.

Insults to the developing brain often result in irreparable damage resulting in long-term deficits in motor and cognitive functions. The only treatment today for hypoxic-ischemic encephalopathy (HIE) in newborns is hypothermia, which has limited clinical benefit. We have studied changes to the blood-brain barriers (BBB) as well as regional cerebral blood flow (rCBF) in a neonatal model of HIE to further understand the underlying pathologic mechanisms. Nine-day old mice pups, brain roughly equivalent to the near-term human fetus, were subjected to hypoxia-ischemia. Hypoxia-ischemia increased BBB permeability to small and large molecules within hours after the insult, which normalized in the following days. The opening of the BBB was associated with changes to BBB protein expression whereas gene transcript levels were increased showing direct molecular damage to the BBB but also suggesting compensatory mechanisms. Brain pathology was closely related to reductions in rCBF during the hypoxia as well as the areas with compromised BBB showing that these are intimately linked. The transient opening of the BBB after the insult is likely to contribute to the pathology but at the same time provides an opportunity for therapeutics to better reach the infarcted areas in the brain.

NRF2-regulation in brain health and disease: implication of cerebral inflammation.

The nuclear factor erythroid 2 related factor 2 (NRF2) is a key regulator of endogenous inducible defense systems in the body. Under physiological conditions NRF2 is mainly located in the cytoplasm. However, in response to oxidative stress, NRF2 translocates to the nucleus and binds to specific DNA sites termed "anti-oxidant response elements" or "electrophile response elements" to initiate transcription of cytoprotective genes. Acute oxidative stress to the brain, such as stroke and traumatic brain injury is increased in animals that are deficient in NRF2. Insufficient NRF2 activation in humans has been linked to chronic diseases such as Parkinson's disease, Alzheimer's disease and amyotrophic lateral sclerosis. New findings have also linked activation of the NRF2 system to anti-inflammatory effects via interactions with NF-κB. Here we review literature on cellular mechanisms of NRF2 regulation, how to maintain and restore NRF2 function and the relationship between NRF2 regulation and brain damage. We bring forward the hypothesis that inflammation via prolonged activation of key kinases (p38 and GSK-3ß) and activation of histone deacetylases gives rise to dysregulation of the NRF2 system in the brain, which contributes to oxidative stress and injury.

Implementing a writing course in an online RN-BSN program.

Scholarly writing is an essential skill for nurses to communicate new research and evidence. Written communication directly relates to patient safety and quality of care. However, few online RN-BSN programs integrate writing instruction into their curricula. Nurses traditionally learn how to write from instructor feedback and often not until midway into their baccalaureate education. Innovative strategies are needed to help nurses apply critical thinking skills to writing. The authors discuss a collaborative project between nursing faculty and technical communication faculty to develop and implement a writing course that is 1 of the 1st courses the students take in the online RN-BSN program.

Involvement of peroxisome proliferator-activated receptor ß/δ (PPAR ß/δ) in BDNF signaling during aging and in Alzheimer disease: possible role of 4-hydroxynonenal (4-HNE).

Aging and many neurological disorders, such as AD, are linked to oxidative stress, which is considered the common effector of the cascade of degenerative events. In this phenomenon, reactive oxygen species play a fundamental role in the oxidative decomposition of polyunsaturated fatty acids, resulting in the formation of a complex mixture of aldehydic end products, such as malondialdehyde, 4-hydroxynonenal, and other alkenals. Interestingly, 4-HNE has been indicated as an intracellular agonist of peroxisome proliferator-activated receptor ß/δ. In this study, we examined, at early and advanced AD stages (3, 9, and 18 months), the pattern of 4-HNE and its catabolic enzyme glutathione S-transferase P1 in relation to the expression of PPARß/δ, BDNF signaling, as mRNA and protein, as well as on their pathological forms (i.e., precursors or truncated forms). The data obtained indicate a novel detrimental age-dependent role of PPAR ß/δ in AD by increasing pro-BDNF and decreasing BDNF/TrkB survival pathways, thus pointing toward the possibility that a specific PPARß/δ antagonist may be used to counteract the disease progression.

Antibody-conjugated PEGylated cerium oxide nanoparticles for specific targeting of Aß aggregates modulate neuronal survival pathways.

Oxidative stress has been found to be associated with the progression of neurodegenerative diseases such as Alzheimer's, Parkinson's, Lou Gehrig's, etc. In the recent years, cerium oxide nanoparticles (CNPs) have been studied as potent antioxidant agents able to exert neuroprotective effects. This work reports polyethylene glycol (PEG)-coated and antibody-conjugated CNPs for the selective delivering to Aß aggregates, and the protective effect against oxidative stress/Aß-mediated neurodegeneration. In this study PEG-coated and anti-Aß antibody-conjugated antioxidant nanoparticles (Aß-CNPs-PEG) were developed, and their effects on neuronal survival and brain-derived neurotrophic factor (BDNF) signaling pathway were examined. Aß-CNPs-PEG specifically targets the Aß aggregates, and concomitant rescue of neuronal survival better than Aß-CNPs, by modulating the BDNF signaling pathway. This proof of concept work may allow in the future, once validated in vivo, for the selective delivery of CNPs only to affected brain areas.

Systemic stimulation of TLR2 impairs neonatal mouse brain development.

BACKGROUND: Inflammation is associated with perinatal brain injury but the underlying mechanisms are not completely characterized. Stimulation of Toll-like receptors (TLRs) through specific agonists induces inflammatory responses that trigger both innate and adaptive immune responses. The impact of engagement of TLR2 signaling pathways on the neonatal brain is still unclear. The aim of this study was to investigate the potential effect of a TLR2 agonist on neonatal brain development. METHODOLOGY/PRINCIPAL FINDINGS: Mice were injected intraperitoneally (i.p.) once a day from postnatal day (PND) 3 to PND11 with endotoxin-free saline, a TLR2 agonist Pam(3)CSK(4) (5 mg/kg) or Lipopolysaccharide (LPS, 0.3 mg/kg). Pups were sacrificed at PND12 or PND53 and brain, spleen and liver were collected and weighed. Brain sections were stained for brain injury markers. Long-term effects on memory function were assessed using the Trace Fear Conditioning test at PND50. After 9 days of Pam(3)CSK(4) administration, we found a decreased volume of cerebral gray matter, white matter in the forebrain and cerebellar molecular layer that was accompanied by an increase in spleen and liver weight at PND12. Such effects were not observed in Pam3CSK4-treated TLR 2-deficient mice. Pam3CSK4-treated mice also displayed decreased hippocampus neuronal density, and increased cerebral microglia density, while there was no effect on caspase-3 or general cell proliferation at PND12. Significantly elevated levels of IL-1ß, IL-6, KC, and MCP-1 were detected after the first Pam3CSK4 injection in brain homogenates of PND3 mice. Pam(3)CSK(4) administration did not affect long-term memory function nor the volume of gray or white matter. CONCLUSIONS/SIGNIFICANCE: Repeated systemic exposure to the TLR2 agonist Pam(3)CSK(4) can have a short-term negative impact on the neonatal mouse brain.

PPARs in Human Neuroepithelial Tumors: PPAR Ligands as Anticancer Therapies for the Most Common Human Neuroepithelial Tumors.

Neuroepithelial tumors represent a heterogeneous class of human tumors including benignant and malignant tumors. The incidence of central nervous system neoplasms ranges from 3.8 to 5.1 cases per 100,000 in the population. Among malignant neuroepithelial tumors, with regard to PPAR ligands, the most extensively studied were tumors of astrocytic origin and neuroblastoma. PPARs are expressed in developing and adult neuroepithelial cells, even if with different localization and relative abundance. The majority of malignant neuroepithelial tumors have poor prognosis and do not respond to conventional therapeutic protocols, therefore, new therapeutic approaches are needed. Natural and synthetic PPAR ligands may represent a starting point for the formulation of new therapeutic approaches to be used as coadjuvants to the standard therapeutic protocols. This review will focus on the major studies dealing with PPAR expression in gliomas and neuroblastoma and the therapeutic implications of using PPAR agonists for the treatment of these neoplasms.

Early biochemical and morphological modifications in the brain of a transgenic mouse model of Alzheimer's disease: a role for peroxisomes.

The central role of peroxisomes in reactive oxygen species and lipid metabolism and their importance in brain functioning are well established. The aim of this work has been to study the peroxisomal population in the Tg2576 mouse model of Alzheimer's disease (AD), at the age of three months when no apparent signs of behavioral, neuroanatomical, cytological, or biochemical alterations have been so far described. The expression and localization of peroxisomal (PMP70, CAT, AOX, and THL) and peroxisome-related proteins (PEX5p, GPX1, SOD1, and SOD2) were studied in the neocortex and hippocampus of transgenic and wild-type animals. Oxidative stress markers (TBARS, acrolein, and 8-OHG) were also evaluated. Our results demonstrate that significant alterations are already detectable at this early stage of the disease and also involve peroxisomes. Their number and protein composition change concomitantly with early oxidative stress. Interestingly, the neocortex shows a compensatory response, consisting in an increase of reactive oxygen species scavenging enzymes, while the hippocampus appears more prone to the oxidative insult. This different behavior could be related to metabolic differences in the two brain areas, also involving peroxisome abundance and/or enzymatic content.