In Vitro Biocompatibility and Endothelial Permeability of Branched Polyglycidols Generated by Ring-Opening Polymerization of Glycidol with B(C6F5)3 under Dry and Wet Conditions.

Polyglycidol or polyglycerol (PG), a polyether widely used in biomedical applications, has not been extensively studied in its branched cyclic form (bcPG), despite extensive research on hyperbranched PG (HPG). This study explores the biomedical promise of bcPG, particularly its ability to cross the blood-brain barrier (BBB). We evaluate in vitro biocompatibility, endothelial permeability, and formation of branched linear PG (blPG) as topological impurities in the presence of water. Small angle X-ray scattering in solution revealed a fractal dimension of approximately two for bcPG and the mixture bc+blPG, suggesting random branching. Comparisons of cytotoxicity and endothelial permeability between bcPG, bc+blPG, and HPG in a BBB model using hCMEC/D3 cells showed different biocompatibility profiles and higher endothelial permeability for HPG. bcPG showed a tendency to accumulate around cell nuclei, in contrast to the behavior of HPG. This study contributes to the understanding of the influence of polymer topology on biological behavior.

Degradable Alternating Copolymers by Radical Copolymerization of 2-Methylen-1,3-dioxepane and Crotonate Esters.

Producing backbone degradable copolymers via free-radical copolymerization is a promising, yet challenging method to develop more sustainable materials for many applications. In this work, we present the copolymerization of 2-methylen-1,3-dioxepane (MDO) with crotonic acid derivative esters. MDO can copolymerize by radical ring-opening polymerization incorporating degradable ester moieties in the polymer backbone, although this can often be difficult due to the very unfavorable reactivity ratios. Crotonic acid derivatives, on the other hand, can be easily produced completely from biomass but are typically very difficult to (co)polymerize due to low propagation rates and very unfavorable reactivity ratios. Herein, we present the surprisingly easy copolymerization between MDO and butyl crotonate (BCr), which shows the ability to form alternating copolymers. The alternating nature of the copolymer was characterized by MALDI-TOF and supported by the reactivity ratios calculated experimentally (rMDO = 0.105 and rBCr = 0.017). The alternating nature of the copolymers favored the degradability that could be achieved under basic conditions (in 2 h, all chains have molar masses smaller than 2 kg/mol). Last, the work was expanded to other crotonate monomers to expand the portfolio and show the potential of this copolymer family.

Synthesis and Characterization of a Luminescent Cyclic Poly(ethylene oxide)-Polypyridyl Ruthenium Complex.

We report the synthesis of a macrocyclic poly(ethylene oxide) (PEO) connected by one [Ru(bpy)3]2+ unit (where bpy = 2,2'-bipyridine), a photoactive metal complex that provides photosensitivity and potential biomedical applications to this polymer structure. The PEO chain provides biocompatibility, water solubility, and topological play. The macrocycles were successfully synthesized by copper-free click cycloaddition between a bifunctional dibenzocyclooctyne (DBCO)-PEO precursor and 4,4'-diazido-2,2'-bipyridine, followed by complexation with [Ru(bpy)2Cl2]. The cyclic product accumulated efficiently in MCF7 cancer cells and exhibited a longer fluorescence lifetime than its linear analogue, likely due to differences in the accessibility of the ligand-centered/intraligand states of Ru polypyridyls in both topologies.

Modulation of Neurolipid Signaling and Specific Lipid Species in the Triple Transgenic Mouse Model of Alzheimer's Disease.

Alzheimer's disease (AD) is a progressive neurodegenerative disorder and the most common cause of dementia in aging populations. Recently, the regulation of neurolipid-mediated signaling and cerebral lipid species was shown in AD patients. The triple transgenic mouse model (3xTg-AD), harboring ßAPPSwe, PS1M146V, and tauP301L transgenes, mimics many critical aspects of AD neuropathology and progressively develops neuropathological markers. Thus, in the present study, 3xTg-AD mice have been used to test the involvement of the neurolipid-based signaling by endocannabinoids (eCB), lysophosphatidic acid (LPA), and sphingosine 1-phosphate (S1P) in relation to the lipid deregulation. [35S]GTPγS autoradiography was used in the presence of specific agonists WIN55,212-2, LPA and CYM5442, to measure the activity mediated by CB1, LPA1, and S1P1 Gi/0 coupled receptors, respectively. Consecutive slides were used to analyze the relative intensities of multiple lipid species by MALDI Mass spectrometry imaging (MSI) with microscopic anatomical resolution. The quantitative analysis of the astrocyte population was performed by immunohistochemistry. CB1 receptor activity was decreased in the amygdala and motor cortex of 3xTg-AD mice, but LPA1 activity was increased in the corpus callosum, motor cortex, hippocampal CA1 area, and striatum. Conversely, S1P1 activity was reduced in hippocampal areas. Moreover, the observed modifications on PC, PA, SM, and PI intensities in different brain areas depend on their fatty acid composition, including decrease of polyunsaturated fatty acid (PUFA) phospholipids and increase of species containing saturated fatty acids (SFA). The regulation of some lipid species in specific brain regions together with the modulation of the eCB, LPA, and S1P signaling in 3xTg-AD mice indicate a neuroprotective adaptation to improve neurotransmission, relieve the myelination dysfunction, and to attenuate astrocyte-mediated neuroinflammation. These results could contribute to identify new therapeutic strategies based on the regulation of the lipid signaling in familial AD patients.

Specific Phospholipid Modulation by Muscarinic Signaling in a Rat Lesion Model of Alzheimer's Disease.

Alzheimer's disease (AD) represents the most common cause of dementia worldwide and has been consistently associated with the loss of basal forebrain cholinergic neurons (BFCNs) leading to impaired cholinergic neurotransmission, aberrant synaptic function, and altered structural lipid metabolism. In this sense, membrane phospholipids (PLs) can be used for de novo synthesis of choline (Ch) for the further obtaining of acetylcholine (ACh) when its availability is compromised. Specific lipid species involved in the metabolism of Ch have been identified as possible biomarkers of phenoconversion to AD. Using a rat model of BFCN lesion, we have evaluated the lipid composition and muscarinic signaling in brain areas related to cognitive processes. The loss of BFCN resulted in alterations of varied lipid species related to Ch metabolism at nucleus basalis magnocellularis (NMB) and cortical projection areas. The activity of muscarinic receptors (mAChR) was decreased in the NMB and increased in the hippocampus according to the subcellular distribution of M1/M2 mAChR which could explain the learning and memory impairment reported in this AD rat model. These results suggest that the modulation of specific lipid metabolic routes could represent an alternative therapeutic strategy to potentiate cholinergic neurotransmission and preserve cell membrane integrity in AD.

A green synthesis of nanocatalysts based on reduced graphene oxide/magnetic nanoparticles for the degradation of Acid Red 1.

The increasing amount of organic dye-polluted wastewater from the textile industry makes the development of techniques for the efficient purification and reuse of wastewater an urgent issue. Accordingly, solid adsorbents based on three-dimensional (3D) reduced graphene oxide (rGO) aerogels combined with magnetic nanoparticles (rGO@Fe3O4) appear to be potential materials, which offer fast and efficient discoloration of dye solutions by dye adsorption, simultaneously acting as Fenton reaction nanocatalysts, and thus may eliminate organic dyes. In this work, 3D rGO@Fe3O4 aerogel nanocatalysts were synthesized via a low-energy, simple, one-step in situ method, in which GO and FeSO4·7H2O were simultaneously reduced. Consequently, monolithic porous nanocatalyst 3D structures were obtained, with a specific surface area of 241 m2 g-1 and pore volume 0.39 cm3 g-1. The nanocatalysts were applied for the degradation of Acid Red 1 azo-dye in aqueous solution in the presence of hydrogen peroxide, without the need for external energy. The effect of the adsorbent dose, and concentration of dye and peroxide on the dye removal was studied. The degradation of the dye was monitored by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. It was found that an increase in the amount of peroxide allowed complete degradation of the dye together with high molar mass side-products with a conjugated aromatic structure. The good nanocatalyst performance was explained based on the charge-transfer complex established between rGO and the magnetic nanoparticles, allowing the regeneration of ferrous ions during the Fenton process.

CB1 and LPA1 Receptors Relationship in the Mouse Central Nervous System.

Neurolipids are a class of bioactive lipids that are produced locally through specific biosynthetic pathways in response to extracellular stimuli. Neurolipids are important endogenous regulators of neural cell proliferation, differentiation, oxidative stress, inflammation and apoptosis. Endocannabinoids (eCBs) and lysophosphatidic acid (LPA) are examples of this type of molecule and are involved in neuroprotection. The present study analyzes a possible relationship of the main receptor subtypes for both neurolipid systems that are present in the central nervous system, the CB1 and LPA1 receptors, by using brain slices from CB1 KO mice and LPA1-null mice. Receptor-mediated G protein activation and glycerophospholipid regulation of potential precursors of their endogenous neurotransmitters were measured by two different in vitro imaging techniques, functional autoradiography and imaging mass spectrometry (IMS), respectively. Possible crosstalk between CB1 and LPA1 receptors was identified in specific areas of the brain, such as the amygdala, where LPA1 receptor activity is upregulated in CB1 KO mice. More evidence of an interaction between both systems was that the CB1-mediated activity was clearly increased in the prefrontal cortex and cerebellum of LPA1-null mice. The eCB system was specifically over-activated in regions where LPA1 has an important signaling role during embryonic development. The modifications on phospholipids (PLs) observed in these genetically modified mice by using the IMS technique indicated the regulation of some of the PL precursors of both LPA and eCBs in specific brain areas. For example, phosphatidylcholine (PC) (36:1) was detected as a potential LPA precursor, and phosphatidylethanolamine (PE) (40:6) and PE (p18:0/22:6) as potential eCB precursors. The absence of the main cerebral receptors for LPA or eCB systems is able to induce modulation on the other at the levels of both signaling and synthesis of endogenous neurotransmitters, indicating adaptive responses between both systems during prenatal and/or postnatal development.

Molecular composition of the human primary visual cortex profiled by multimodal mass spectrometry imaging.

The primary visual cortex (area V1) is an extensively studied part of the cerebral cortex with well-characterized connectivity, cellular and molecular architecture and functions (for recent reviews see Amunts and Zilles, Neuron 88:1086-1107, 2015; Casagrande and Xu, Parallel visual pathways: a comparative perspective. The visual neurosciences, MIT Press, Cambridge, pp 494-506, 2004). In humans, V1 is defined by heavily myelinated fibers arriving from the radiatio optica that form the Gennari stripe in cortical layer IV, which is further subdivided into laminae IVa, IVb, IVcα and IVcß. Due to this unique laminar pattern, V1 represents an excellent region to test whether multimodal mass spectrometric imaging could reveal novel biomolecular markers for a functionally relevant parcellation of the human cerebral cortex. Here we analyzed histological sections of three post-mortem brains with matrix-assisted laser desorption/ionization mass spectrometry imaging and laser ablation inductively coupled plasma mass spectrometry imaging to investigate the distribution of lipids, proteins and metals in human V1. We identified 71 peptides of 13 different proteins by in situ tandem mass spectrometry, of which 5 proteins show a differential laminar distribution pattern revealing the border between V1 and V2. High-accuracy mass measurements identified 123 lipid species, including glycerolipids, glycerophospholipids and sphingolipids, of which at least 20 showed differential distribution within V1 and V2. Specific lipids labeled not only myelinated layer IVb, but also IVa and especially IVc in a layer-specific manner, but also and clearly separated V1 from V2. Elemental imaging further showed a specific accumulation of copper in layer IV. In conclusion, multimodal mass spectrometry imaging identified novel biomolecular and elemental markers with specific laminar and inter-areal differences. We conclude that mass spectrometry imaging provides a promising new approach toward multimodal, molecule-based cortical parcellation.

Anatomical location of LPA1 activation and LPA phospholipid precursors in rodent and human brain.

Lysophosphatidic acid (LPA) is a signaling molecule that binds to six known G protein-coupled receptors: LPA1 -LPA6 . LPA evokes several responses in the CNS, including cortical development and folding, growth of the axonal cone and its retraction process. Those cell processes involve survival, migration, adhesion proliferation, differentiation, and myelination. The anatomical localization of LPA1 is incompletely understood, particularly with regard to LPA binding. Therefore, we have used functional [(35) S]GTPγS autoradiography to verify the anatomical distribution of LPA1 binding sites in adult rodent and human brain. The greatest activity was observed in myelinated areas of the white matter such as corpus callosum, internal capsule and cerebellum. MaLPA1 -null mice (a variant of LPA1 -null) lack [(35) S]GTPγS basal binding in white matter areas, where the LPA1 receptor is expressed at high levels, suggesting a relevant role of the activity of this receptor in the most myelinated brain areas. In addition, phospholipid precursors of LPA were localized by MALDI-IMS in both rodent and human brain slices identifying numerous species of phosphatides and phosphatidylcholines. Both phosphatides and phosphatidylcholines species represent potential LPA precursors. The anatomical distribution of these precursors in rodent and human brain may indicate a metabolic relationship between LPA and LPA1 receptors. Lysophosphatidic acid (LPA) is a signaling molecule that binds to six known G protein-coupled receptors (GPCR), LPA1 to LPA6 . LPA evokes several responses in the central nervous system (CNS), including cortical development and folding, growth of the axonal cone and its retraction process. We used functional [(35) S]GTPγS autoradiography to verify the anatomical distribution of LPA1 -binding sites in adult rodent and human brain. The distribution of LPA1 receptors in rat, mouse and human brains show the highest activity in white matter myelinated areas. The basal and LPA-evoked activities are abolished in MaLPA1 -null mice. The phospholipid precursors of LPA are localized by MALDI-IMS. The anatomical distribution of LPA precursors in rodent and human brain suggests a relationship with functional LPA1 receptors.

Neurotransmitter receptor localization: from autoradiography to imaging mass spectrometry.

Autoradiography is used to determine the anatomical distribution of biological molecules in human tissue and experimental animal models. This method is based on the analysis of the specific binding of radiolabeled compounds to locate neurotransmitter receptors or transporters in fresh frozen tissue slices. The anatomical resolution obtained by quantification of the radioligands has allowed the density of receptor proteins to be mapped over the last 40 years. The data yielded by autoradiography identify the receptors at their specific microscopic localization in the tissues and also in their native microenvironment, the intact cell membrane. Furthermore, in functional autoradiography, the effects of small molecules on the activity of G protein-coupled receptors are evaluated. More recently, autoradiography has been combined with membrane microarrays to improve the high-throughput screening of compounds. These technical advances have made autoradiography an essential analytical method for the progress of drug discovery. We include the future prospects and some preliminary results for imaging mass spectrometry (IMS) as a useful new method in pharmacodynamic and pharmacokinetic studies, complementing autoradiographic studies. IMS results could also be presented as density maps of molecules, proteins, and metabolites in tissue sections that can be identified, localized, and quantified, with the advantage of avoiding any labeling of marker molecules. The limitations and future developments of these techniques are discussed here.

Type-1 cannabinoid receptor activity during Alzheimer's disease progression.

The activity of CB1 cannabinoid receptors was studied in postmortem brain samples of Alzheimer's disease (AD) patients during clinical deterioration. CB1 activity was higher at earlier AD stages in limited hippocampal areas and internal layers of frontal cortex, but a decrease was observed at the advanced stages. The pattern of modification appears to indicate initial hyperactivity of the endocannabinoid system in brain areas that lack classical histopathological markers at earlier stages of AD, indicating an attempt to compensate for the initial synaptic impairment, which is then surpassed by disease progression. These results suggest that initial CB1 stimulation might have therapeutic relevance.