Solution Dynamics of Covalent Open-[60]Fullerene Dimers.

The translational diffusivity of covalent open-[60]fullerene dimers in an organic solvent was found to be well describable by a prolate ellipsoid model while a monomeric open-[60]fullerene behaves like a sphere model. The water association dynamics were examined for two open-[60]fullerene dimers, showing a higher water affinity for the sp3-linked dimer relative to sp2-linked dimer owing to an effective orbital-orbital overlap identified by π(fullerene)→σ*(H2O) interactions as suggested by theoretical calculations.

Phosphine-Mediated Dimerization of Open-[60]Fullerenes.

By a reaction of trimethylphosphine with an open-[60]fullerene, corresponding dimers could be generated via two-fold deoxygenation processes even though the formation of ß-oxo-phosphorous ylide is inevitable, a part of which is hydrolyzed to yield an α-methylene carbonyl derivative. Nevertheless, Wittig reaction and aldol condensation did not proceed well, indicating the presence of an unknown dimerization pathway. In the ylide formation, 1-phosphonium-3-carbabetaine was previously proposed as a key intermediate. Upon assuming that the betaine also participates in the dimerization process, we examined a possible reaction pathway computationally. As the results, the betaine formed by a reaction with the first phosphine was suggested to undergo nucleophilic addition to an unreacted molecule of the open-[60]fullerene, yielding an epoxide dimer which is then deoxygenated by the second phosphine to furnish the desired open-[60]fullerene dimer.

Synthesis of inter-[60]fullerene conjugates with inherent chirality.

Coalescence of [60]fullerenes potentially produces hypothetical nanocarbon assemblies with non-naturally occurring topologies. Since the discovery of [60]fullerene in 1985, coalesced [60]fullerene oligomers have only been observed as transient species by transmission electron microscopy during an oligomerization process under a high electron acceleration voltage. Herein, we showcase the rational synthesis of covalent assemblies consisting of inherently chiral open-[60]fullerenes. The crystallographic analyses unveiled double-caged structures of non-conjugated and conjugated inter-[60]fullerene hybrids, in which the two [60]fullerene cages are bounds to each other through a covalent linkage. The former one further assembles via a heterochiral recognition so that four carbon cages are arranged in a tetrahedral manner both in solution and solid state. Reflecting radially-conjugated double π-surface nature, the inter-[60]fullerene conjugate exhibits strong electronic communication in its reduced states, intense absorption behavior, and chiroptical activity with a dissymmetry factor of 0.21 (at 674 nm) which breaks the record for known chiral organic molecules.

[A Case of Prostate Cancer Diagnosed by the Bone Biopsy of the Right Iliac Metastatic Lesion].

A 73-year-old man was referred to our hospital because of a high prostate specific antigen (PSA) level. The PSA level at our hospital was 63.5 ng/ml. Pelvic magnetic resonance imaging (MRI) showed findings strongly suggestive of multiple pelvic bone metastases, but no obvious malignant findings in the prostate. A 12-core prostate biopsy was performed and no cancer was detected. Computed tomography and bone scintigraphy showed findings suspicious of bone metastases in the sternum, thoracolumbar spine, pelvic bone, and sacrum. Spine MRI revealed a mass in the vertebral body from the eighth thoracic vertebra to the first lumbar vertebra. A biopsy of the right iliac crest showed adenocarcinoma and was positive for PSA staining, leading to the diagnosis of multiple bone metastases of prostate cancer. Abiraterone acetate in combination with androgen deprivation was started. He received medication and radiation therapy to his sternum for pain relief. Spine MRI after 4 months showed decreased vertebral body weights and serum PSA levels were ＜0.003 ng/ml after 5 months. Seventeen months after treatment, PSA remains below 0.003 ng/ml, and the patient is currently pain-free.

Near-Infrared-Absorbing Chiral Open [60]Fullerenes.

Though [60]fullerene is an achiral molecular nanocarbon with Ih symmetry, it could attain an inherent chirality depending upon a functionalization pattern. The conventional chiral induction of C60 relies mainly upon a multiple addition affording a mixture of achiral and chiral isomers while their chiral function would be largely offset by the existence of pseudo-mirror plane(s). These are major obstacles to proceed further study on fullerene chirality and yet leave its understanding elusive. Herein, we showcase a carbene-mediated synthesis of C1 -symmetric chiral open [60]fullerenes showing an intense far-red to near-infrared absorption. The large dissymmetry factor of |gabs |=0.12 was achieved at λ=820 nm for circular dichroism in benzonitrile. This is, in general, unachievable by other small chiral organic molecules, demonstrating the potential usage of open [60]fullerenes as novel types of chiral chromophores.

Phosphorus ylides of cage-opened sulphide [60]fullerene derivatives.

The partial structural change in cage-opened C60 derivatives by replacing a carbonyl group with a sulphide moiety significantly lowers the chemical stability of the corresponding phosphorus ylides. Thus they readily undergo in situ hydrolysis to give an α-methylene carbonyl compound. Owing to the elevated LUMO level, the near-infrared absorption band was hypsochromically shifted from λedge 880 to 800 nm.

Chiral Open-[60]Fullerene Ligands with Giant Dissymmetry Factors.

The optical resolution of open-[60]fullerenes has been limited to only one example since 1998, while the recent advances revealed the excellence of fullerenes as revisited chiral functional materials. Different from conventional chiral induction on [60]fullerene by a multiple-functionalization, a random disruption of the spherical π-conjugation is avoidable for open-[60]fullerenes. Moreover, the macrocyclic orifices enable a metal coordination which endows modulated electronic structures on chiral chromophores. Herein, we showcase Li+-coordination behavior and optical resolution of three chiral open-[60]fullerene ligands, showing a giant dissymmetry factor up to 0.20 owing to a congenital topology of the spherical π-conjugation.

Reactions on a 1,2-Dicarbonyl Moiety of a Fullerene Skeleton.

A 1,2-dicarbonyl moiety on a cage-opened fullerene skeleton is one of suitable building blocks for the further derivatization. Herein, we discuss the chemical transformation of a 1,2-dicarbonyl compound into ß-oxo-phosphorus ylide, acid anhydride, and α-methylene carbonyl derivatives. Despite possessing a sterically small methylene unit in the last one, the release of an encapsulated water molecule was significantly supressed whereas the ß-oxo-phosphorus ylide bearing three bulky p-tolyl groups on the P-atom enabled the faster insertion/release dynamics, implying the flexibility of the phosphonium substituent. The replacement of the carbonyl group with phosphorus ylide and methylene units largely varied electrochemical properties of the fullerene skeleton, likely arising from the anionic charge delocalized over the entire molecule and removal of an electron-withdrawable carbonyl group, respectively.

Nonclassical Abramov Products Formed on Orifices of Cage-Opened C60 Derivatives.

By nucleophilic addition of phosphite P(OMe)3 to a cage-opened C60 derivative, α-hydrophosphate and enol phosphate were obtained as kinetic and thermodynamic products, respectively. Different from classical Abramov products bearing a phosphorus-carbon bond, these products have a phosphorus-oxygen bond. The observed anomaly originates from the fully conjugated π system, which significantly stabilizes zwitterionic intermediates bearing a phosphorus-oxygen bond. The thus formed enol phosphate was found to exhibit an intense absorption band that extended to 730 nm, reflecting the intramolecular charge-transfer transitions. We also report domino phosphorylation reactions, which gave a cage-opened C60 derivative bearing a direct P-C bond.

Organophosphorus zwitterions engaged in a conjugated macrocycle on fullerene.

Organophosphorus zwitterions are one of the most important but elusive intermediates for carbon-carbon bond formation in synthetic chemistry and biology. However, a lack of isolated examples due to their lability has hampered in-depth understanding of structures and their reaction mechanisms. In this study, we crystallographically reveal the solid-state structure of a phosha-Michael adduct engaged in a cage-opened C60 skeleton, which is formed as a kinetic product. This compound exhibits dark brown colour in solution with an intense absorption band that extends to 1000 nm, reflecting intramolecular charge transfer transitions. From the 1,2-dicarbonyl moiety on the conjugated orifice, ß-oxo-phosphorus ylide is formed as a thermodynamic product. The reaction mechanism that has long been disputed is examined by experimental and theoretical studies, showing a pathway which includes an SN2 reaction as a key step instead of the hitherto considered carbene pathway.

NitroSynapsin for the treatment of neurological manifestations of tuberous sclerosis complex in a rodent model.

Tuberous sclerosis (TSC) is an autosomal dominant disorder caused by heterozygous mutations in the TSC1 or TSC2 gene. TSC is often associated with neurological, cognitive, and behavioral deficits. TSC patients also express co-morbidity with anxiety and mood disorders. The mechanism of pathogenesis in TSC is not entirely clear, but TSC-related neurological symptoms are accompanied by excessive glutamatergic activity and altered synaptic spine structures. To address whether extrasynaptic (e)NMDA-type glutamate receptor (NMDAR) antagonists, as opposed to antagonists that block physiological phasic synaptic activity, can ameliorate the synaptic and behavioral features of this disease, we utilized the Tsc2+/- mouse model of TSC to measure biochemical, electrophysiological, histological, and behavioral parameters in the mice. We found that antagonists that preferentially block tonic activity as found at eNMDARs, particularly the newer drug NitroSynapsin, provide biological and statistically significant improvement in Tsc2+/- phenotypes. Accompanying this improvement was correction of activity in the p38 MAPK-TSC-Rheb-mTORC1-S6K1 pathway. Deficits in hippocampal long-term potentiation (LTP), histological loss of synapses, and behavioral fear conditioning in Tsc2+/- mice were all improved after treatment with NitroSynapsin. Taken together, these results suggest that amelioration of excessive excitation, by limiting aberrant eNMDAR activity, may represent a novel treatment approach for TSC.

Molecular Pathway to Protection From Age-Dependent Photoreceptor Degeneration in Mef2 Deficiency.

Purpose: Photoreceptor degeneration in the retina is a major cause of blindness in humans. Elucidating mechanisms of degenerative and neuroprotective pathways in photoreceptors should afford identification and development of therapeutic strategies. Methods: We used mouse genetic models and improved methods for retinal explant cultures. Retinas were enucleated from Mef2d+/+ and Mef2d-/- mice, stained for MEF2 proteins and outer nuclear layer thickness, and assayed for apoptotic cells. Chromatin immunoprecipitation (ChIP) assays revealed MEF2 binding, and RT-qPCR showed levels of transcription factors. We used AAV2 and electroporation to express genes in retinal explants and electroretinograms to assess photoreceptor functionality. Results: We identify a prosurvival MEF2D-PGC1α pathway that plays a neuroprotective role in photoreceptors. We demonstrate that Mef2d-/- mouse retinas manifest decreased expression of PGC1α and increased photoreceptor cell loss, resulting in the absence of light responses. Molecular repletion of PGC1α protects Mef2d-/- photoreceptors and preserves light responsivity. Conclusions: These results suggest that the MEF2-PGC1α cascade may represent a new therapeutic target for drugs designed to protect photoreceptors from developmental- and age-dependent loss.

MEF2D haploinsufficiency downregulates the NRF2 pathway and renders photoreceptors susceptible to light-induced oxidative stress.

Gaining mechanistic insight into interaction between causative factors of complex multifactorial diseases involving photoreceptor damage might aid in devising effective therapies. Oxidative stress is one of the potential unifying mechanisms for interplay between genetic and environmental factors that contribute to photoreceptor pathology. Interestingly, the transcription factor myocyte enhancer factor 2d (MEF2D) is known to be important in photoreceptor survival, as knockout of this transcription factor results in loss of photoreceptors in mice. Here, using a mild light-induced retinal degeneration model, we show that the diminished MEF2D transcriptional activity in Mef2d+/- retina is further reduced under photostimulation-induced oxidative stress. Reactive oxygen species cause an aberrant redox modification on MEF2D, consequently inhibiting transcription of its downstream target, nuclear factor (erythroid-derived 2)-like 2 (NRF2). NRF2 is a master regulator of phase II antiinflammatory and antioxidant gene expression. In the Mef2d heterozygous mouse retina, NRF2 is not up-regulated to a normal degree in the face of light-induced oxidative stress, contributing to accelerated photoreceptor cell death. Furthermore, to combat this injury, we found that activation of the endogenous NRF2 pathway using proelectrophilic drugs rescues photoreceptors from photo-induced oxidative stress and may therefore represent a viable treatment for oxidative stress-induced photoreceptor degeneration, which is thought to contribute to some forms of retinitis pigmentosa and age-related macular degeneration.

Aberrant protein S-nitrosylation contributes to the pathophysiology of neurodegenerative diseases.

Nitric oxide (NO) is a gasotransmitter that impacts fundamental aspects of neuronal function in large measure through S-nitrosylation, a redox reaction that occurs on regulatory cysteine thiol groups. For instance, S-nitrosylation regulates enzymatic activity of target proteins via inhibition of active site cysteine residues or via allosteric regulation of protein structure. During normal brain function, protein S-nitrosylation serves as an important cellular mechanism that modulates a diverse array of physiological processes, including transcriptional activity, synaptic plasticity, and neuronal survival. In contrast, emerging evidence suggests that aging and disease-linked environmental risk factors exacerbate nitrosative stress via excessive production of NO. Consequently, aberrant S-nitrosylation occurs and represents a common pathological feature that contributes to the onset and progression of multiple neurodegenerative disorders, including Alzheimer's, Parkinson's, and Huntington's diseases. In the current review, we highlight recent key findings on aberrant protein S-nitrosylation showing that this reaction triggers protein misfolding, mitochondrial dysfunction, transcriptional dysregulation, synaptic damage, and neuronal injury. Specifically, we discuss the pathological consequences of S-nitrosylated parkin, myocyte enhancer factor 2 (MEF2), dynamin-related protein 1 (Drp1), protein disulfide isomerase (PDI), X-linked inhibitor of apoptosis protein (XIAP), and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) under neurodegenerative conditions. We also speculate that intervention to prevent these aberrant S-nitrosylation events may produce novel therapeutic agents to combat neurodegenerative diseases.

Transcriptional profiling of MEF2-regulated genes in human neural progenitor cells derived from embryonic stem cells.

[Briefly describe the contents of the Data in Brief article. Tell the reader the repository and reference number for the data in the abstract to.] The myocyte enhancer factor 2 (MEF2) family of transcription factors is highly expressed in the brain, and constitutes a key determinant of neuronal survival, differentiation, and synaptic plasticity. However, genome-wide transcriptional profiling of MEF2-regulated genes has not yet been fully elucidated, particularly at the neural stem cell stage. Here we report the results of microarray analysis comparing mRNAs isolated from human neural progenitor/stem cells (hNPCs) derived from embryonic stem cells expressing a control vector versus progenitors expressing a constitutively-active form of MEF2 (MEF2CA), which increases MEF2 activity. Microarray experiments were performed using the Illumina Human HT-12 V4.0 expression beadchip (GEO#: GSE57184). By comparing vector-control cells to MEF2CA cells, microarray analysis identified 1880 unique genes that were differentially expressed. Among these genes, 1121 genes were upregulated and 759 genes were down-regulated. Our results provide a valuable resource for identifying transcriptional targets of MEF2 in hNPCs.

S-Nitrosylation in neurogenesis and neuronal development.

BACKGROUND: Nitric oxide (NO) is a pleiotropic messenger molecule. The multidimensional actions of NO species are, in part, mediated by their redox nature. Oxidative posttranslational modification of cysteine residues to regulate protein function, termed S-nitrosylation, constitutes a major form of redox-based signaling by NO. SCOPE OF REVIEW: S-Nitrosylation directly modifies a number of cytoplasmic and nuclear proteins in neurons. S-Nitrosylation modulates neuronal development by reaction with specific proteins, including the transcription factor MEF2. This review focuses on the impact of S-nitrosylation on neurogenesis and neuronal development. MAJOR CONCLUSIONS: Functional characterization of S-nitrosylated proteins that regulate neuronal development represents a rapidly emerging field. Recent studies reveal that S-nitrosylation-mediated redox signaling plays an important role in several biological processes essential for neuronal differentiation and maturation. GENERAL SIGNIFICANCE: Investigation of S-nitrosylation in the nervous system has elucidated new molecular and cellular mechanisms for neuronal development. S-Nitrosylated proteins in signaling networks modulate key events in brain development. Dysregulation of this redox-signaling pathway may contribute to neurodevelopmental disabilities such as autism spectrum disorder (ASD). Thus, further elucidation of the involvement of S-nitrosylation in brain development may offer potential therapeutic avenues for neurodevelopmental disorders. This article is part of a Special Issue entitled Redox regulation of differentiation and de-differentiation.

Oligomeric Aß-induced synaptic dysfunction in Alzheimer's disease.

Alzheimer's disease (AD) is a devastating disease characterized by synaptic and neuronal loss in the elderly. Compelling evidence suggests that soluble amyloid-ß peptide (Aß) oligomers induce synaptic loss in AD. Aß-induced synaptic dysfunction is dependent on overstimulation of N-methyl-D-aspartate receptors (NMDARs) resulting in aberrant activation of redox-mediated events as well as elevation of cytoplasmic Ca2+, which in turn triggers downstream pathways involving phospho-tau (p-tau), caspases, Cdk5/dynamin-related protein 1 (Drp1), calcineurin/PP2B, PP2A, Gsk-3ß, Fyn, cofilin, and CaMKII and causes endocytosis of AMPA receptors (AMPARs) as well as NMDARs. Dysfunction in these pathways leads to mitochondrial dysfunction, bioenergetic compromise and consequent synaptic dysfunction and loss, impaired long-term potentiation (LTP), and cognitive decline. Evidence also suggests that Aß may, at least in part, mediate these events by causing an aberrant rise in extrasynaptic glutamate levels by inhibiting glutamate uptake or triggering glutamate release from glial cells. Consequent extrasynaptic NMDAR (eNMDAR) overstimulation then results in synaptic dysfunction via the aforementioned pathways. Consistent with this model of Aß-induced synaptic loss, Aß synaptic toxicity can be partially ameliorated by the NMDAR antagonists (such as memantine and NitroMemantine). PSD-95, an important scaffolding protein that regulates synaptic distribution and activity of both NMDA and AMPA receptors, is also functionally disrupted by Aß. PSD-95 dysregulation is likely an important intermediate step in the pathological cascade of events caused by Aß. In summary, Aß-induced synaptic dysfunction is a complicated process involving multiple pathways, components and biological events, and their underlying mechanisms, albeit as yet incompletely understood, may offer hope for new therapeutic avenues.

S-nitrosylation-mediated redox transcriptional switch modulates neurogenesis and neuronal cell death.

Redox-mediated posttranslational modifications represent a molecular switch that controls major mechanisms of cell function. Nitric oxide (NO) can mediate redox reactions via S-nitrosylation, representing transfer of an NO group to a critical protein thiol. NO is known to modulate neurogenesis and neuronal survival in various brain regions in disparate neurodegenerative conditions. However, a unifying molecular mechanism linking these phenomena remains unknown. Here, we report that S-nitrosylation of myocyte enhancer factor 2 (MEF2) transcription factors acts as a redox switch to inhibit both neurogenesis and neuronal survival. Structure-based analysis reveals that MEF2 dimerization creates a pocket, facilitating S-nitrosylation at an evolutionally conserved cysteine residue in the DNA binding domain. S-Nitrosylation disrupts MEF2-DNA binding and transcriptional activity, leading to impaired neurogenesis and survival in vitro and in vivo. Our data define a molecular switch whereby redox-mediated posttranslational modification controls both neurogenesis and neurodegeneration via a single transcriptional signaling cascade.

ATM-dependent phosphorylation of MEF2D promotes neuronal survival after DNA damage.

Mutations in the ataxia telangiectasia mutated (ATM) gene, which encodes a kinase critical for the normal DNA damage response, cause the neurodegenerative disorder ataxia-telangiectasia (AT). The substrates of ATM in the brain are poorly understood. Here we demonstrate that ATM phosphorylates and activates the transcription factor myocyte enhancer factor 2D (MEF2D), which plays a critical role in promoting survival of cerebellar granule cells. ATM associates with MEF2D after DNA damage and phosphorylates the transcription factor at four ATM consensus sites. Knockdown of endogenous MEF2D with a short-hairpin RNA (shRNA) increases sensitivity to etoposide-induced DNA damage and neuronal cell death. Interestingly, substitution of endogenous MEF2D with an shRNA-resistant phosphomimetic MEF2D mutant protects cerebellar granule cells from cell death after DNA damage, whereas an shRNA-resistant nonphosphorylatable MEF2D mutant does not. In vivo, cerebella in Mef2d knock-out mice manifest increased susceptibility to DNA damage. Together, our results show that MEF2D is a substrate for phosphorylation by ATM, thus promoting survival in response to DNA damage. Moreover, dysregulation of the ATM-MEF2D pathway may contribute to neurodegeneration in AT.

Isogenic human iPSC Parkinson's model shows nitrosative stress-induced dysfunction in MEF2-PGC1α transcription.

Parkinson's disease (PD) is characterized by loss of A9 dopaminergic (DA) neurons in the substantia nigra pars compacta (SNpc). An association has been reported between PD and exposure to mitochondrial toxins, including environmental pesticides paraquat, maneb, and rotenone. Here, using a robust, patient-derived stem cell model of PD allowing comparison of A53T α-synuclein (α-syn) mutant cells and isogenic mutation-corrected controls, we identify mitochondrial toxin-induced perturbations in A53T α-syn A9 DA neurons (hNs). We report a pathway whereby basal and toxin-induced nitrosative/oxidative stress results in S-nitrosylation of transcription factor MEF2C in A53T hNs compared to corrected controls. This redox reaction inhibits the MEF2C-PGC1α transcriptional network, contributing to mitochondrial dysfunction and apoptotic cell death. Our data provide mechanistic insight into gene-environmental interaction (GxE) in the pathogenesis of PD. Furthermore, using small-molecule high-throughput screening, we identify the MEF2C-PGC1α pathway as a therapeutic target to combat PD.