In Vivo Methods to Study Uptake of Nanoparticles into the Brain

Several in vivo techniques have been developed to study and measure the uptake of CNS compounds into the brain. With these techniques, various parameters can be determined after drug administration, including the blood-to-brain influx constant (Kin), the permeability-surface area (PS) product, and the brain uptake index (BUI). These techniques have been mostly used for drugs that are expected to enter the brain via transmembrane diffusion or by carrier-mediated transcytosis. Drugs that have limitations in entering the brain via such pathways have been encapsulated in nanoparticles (based on lipids or synthetic polymers) to enhance brain uptake. Nanoparticles are different from CNS compounds in size, composition and uptake mechanisms. This has led to different methods and approaches to study brain uptake in vivo. Here we discuss the techniques generally used to measure nanoparticle uptake in addition to the techniques used for CNS compounds. Techniques include visualization methods, behavioral tests, and quantitative methods

Genome-wide haplotype association study identifies the FRMD4A gene as a risk locus for Alzheimer's disease.

International audienceRecently, several genome wide association studies (GWAS) have led to the discovery of 9 new loci of genetic susceptibility in Alzheimer's disease (AD). However, the landscape of the AD genetic susceptibility is far away to be complete and in addition to single-SNP analyses as performed in conventional GWAS, complementary strategies need to be applied to overcome limitations inherent to this type of approaches.. We performed a genome wide haplotype association (GWHA) study in the EADI1 study (n=2,025 AD cases and 5,328 controls) by applying a sliding-windows approach. After exclusion of loci already known to be involved in AD (APOE, BIN1 and CR1), 91 regions with suggestive haplotype effects were identified. In a second step, we attempted to replicate the best suggestive haplotype associations in the GERAD1 consortium (2,820 AD cases and 6,356 controls) and observed that 9 of them showed nominal association. In a third step, we tested relevant haplotype associations in a combined analysis of five additional case-control studies (5,093 AD cases and 4,061 controls). We consistently replicated the association of a haplotype within FRMD4A on Chr.10p13 in all the data set analysed (OR=1.68, 95% CI 1.43- 1.96; p=1.1x10-10). We finally searched for association between SNPs within the FRMD4A locus and Ab plasma concentrations in three independent non demented populations (n=2,579). We reported that polymorphisms were associated with plasma Ab42/Ab40 ratio (best signal, p=5.4x10-7). In conclusion, combining both GWHA study and a conservative three-stage replication approach, we characterised FRMD4A as a new genetic risk factor of AD

CNS Penetration of Intrathecal-Lumbar Idursulfase in the Monkey, Dog and Mouse: Implications for Neurological Outcomes of Lysosomal Storage Disorder

A major challenge for the treatment of many central nervous system (CNS) disorders is the lack of convenient and effective methods for delivering biological agents to the brain. Mucopolysaccharidosis II (Hunter syndrome) is a rare inherited lysosomal storage disorder resulting from a deficiency of iduronate-2-sulfatase (I2S). I2S is a large, highly glycosylated enzyme. Intravenous administration is not likely to be an effective therapy for disease-related neurological outcomes that require enzyme access to the brain cells, in particular neurons and oligodendrocytes. We demonstrate that intracerebroventricular and lumbar intrathecal administration of recombinant I2S in dogs and nonhuman primates resulted in widespread enzyme distribution in the brain parenchyma, including remarkable deposition in the lysosomes of both neurons and oligodendrocytes. Lumbar intrathecal administration also resulted in enzyme delivery to the spinal cord, whereas little enzyme was detected there after intraventricular administration. Mucopolysaccharidosis II model is available in mice. Lumbar administration of recombinant I2S to enzyme deficient animals reduced the storage of glycosaminoglycans in both superficial and deep brain tissues, with concurrent morphological improvements. The observed patterns of enzyme transport from cerebrospinal fluid to the CNS tissues and the resultant biological activity (a) warrant further investigation of intrathecal delivery of I2S via lumbar catheter as an experimental treatment for the neurological symptoms of Hunter syndrome and (b) may have broader implications for CNS treatment with biopharmaceuticals

Advancing brain barriers RNA sequencing: guidelines from experimental design to publication

Background: RNA sequencing (RNA-Seq) in its varied forms has become an indispensable tool for analyzing differential gene expression and thus characterization of specific tissues. Aiming to understand the brain barriers genetic signature, RNA seq has also been introduced in brain barriers research. This has led to availability of both, bulk and single-cell RNA-Seq datasets over the last few years. If appropriately performed, the RNA-Seq studies provide powerful datasets that allow for significant deepening of knowledge on the molecular mechanisms that establish the brain barriers. However, RNA-Seq studies comprise complex workflows that require to consider many options and variables before, during and after the proper sequencing process.Main body: In the current manuscript, we build on the interdisciplinary experience of the European PhD Training Network BtRAIN (https://www.btrain-2020.eu/) where bioinformaticians and brain barriers researchers collaborated to analyze and establish RNA-Seq datasets on vertebrate brain barriers. The obstacles BtRAIN has identified in this process have been integrated into the present manuscript. It provides guidelines along the entire workflow of brain barriers RNA-Seq studies starting from the overall experimental design to interpretation of results. Focusing on the vertebrate endothelial blood–brain barrier (BBB) and epithelial blood-cerebrospinal-fluid barrier (BCSFB) of the choroid plexus, we provide a step-by-step description of the workflow, highlighting the decisions to be made at each step of the workflow and explaining the strengths and weaknesses of individual choices made. Finally, we propose recommendations for accurate data interpretation and on the information to be included into a publication to ensure appropriate accessibility of the data and reproducibility of the observations by the scientific community.Conclusion: Next generation transcriptomic profiling of the brain barriers provides a novel resource for understanding the development, function and pathology of these barrier cells, which is essential for understanding CNS homeostasis and disease. Continuous advancement and sophistication of RNA-Seq will require interdisciplinary approaches between brain barrier researchers and bioinformaticians as successfully performed in BtRAIN. The present guidelines are built on the BtRAIN interdisciplinary experience and aim to facilitate collaboration of brain barriers researchers with bioinformaticians to advance RNA-Seq study design in the brain barriers community

Analysis of the effects of sex hormone background on the rat choroid plexus transcriptome by cDNA microarrays

The choroid plexus (CP) are highly vascularized branched structures that protrude into the ventricles of the brain, and form a unique interface between the blood and the cerebrospinal fluid (CSF), the blood-CSF barrier, that are the main site of production and secretion of CSF. Sex hormones are widely recognized as neuroprotective agents against several neurodegenerative diseases, and the presence of sex hormones cognate receptors suggest that it may be a target for these hormones. In an effort to provide further insight into the neuroprotective mechanisms triggered by sex hormones we analyzed gene expression differences in the CP of female and male rats subjected to gonadectomy, using microarray technology. In gonadectomized female and male animals, 3045 genes were differentially expressed by 1.5-fold change, compared to sham controls. Analysis of the CP transcriptome showed that the top-five pathways significantly regulated by the sex hormone background are olfactory transduction, taste transduction, metabolism, steroid hormone biosynthesis and circadian rhythm pathways. These results represent the first overview of global expression changes in CP of female and male rats induced by gonadectomy and suggest that sex hormones are implicated in pathways with central roles in CP functions and CSF homeostasis

Multiplicity of cerebrospinal fluid functions: New challenges in health and disease

This review integrates eight aspects of cerebrospinal fluid (CSF) circulatory dynamics: formation rate, pressure, flow, volume, turnover rate, composition, recycling and reabsorption. Novel ways to modulate CSF formation emanate from recent analyses of choroid plexus transcription factors (E2F5), ion transporters (NaHCO3 cotransport), transport enzymes (isoforms of carbonic anhydrase), aquaporin 1 regulation, and plasticity of receptors for fluid-regulating neuropeptides. A greater appreciation of CSF pressure (CSFP) is being generated by fresh insights on peptidergic regulatory servomechanisms, the role of dysfunctional ependyma and circumventricular organs in causing congenital hydrocephalus, and the clinical use of algorithms to delineate CSFP waveforms for diagnostic and prognostic utility. Increasing attention focuses on CSF flow: how it impacts cerebral metabolism and hemodynamics, neural stem cell progression in the subventricular zone, and catabolite/peptide clearance from the CNS. The pathophysiological significance of changes in CSF volume is assessed from the respective viewpoints of hemodynamics (choroid plexus blood flow and pulsatility), hydrodynamics (choroidal hypo- and hypersecretion) and neuroendocrine factors (i.e., coordinated regulation by atrial natriuretic peptide, arginine vasopressin and basic fibroblast growth factor). In aging, normal pressure hydrocephalus and Alzheimer's disease, the expanding CSF space reduces the CSF turnover rate, thus compromising the CSF sink action to clear harmful metabolites (e.g., amyloid) from the CNS. Dwindling CSF dynamics greatly harms the interstitial environment of neurons. Accordingly the altered CSF composition in neurodegenerative diseases and senescence, because of adverse effects on neural processes and cognition, needs more effective clinical management. CSF recycling between subarachnoid space, brain and ventricles promotes interstitial fluid (ISF) convection with both trophic and excretory benefits. Finally, CSF reabsorption via multiple pathways (olfactory and spinal arachnoidal bulk flow) is likely complemented by fluid clearance across capillary walls (aquaporin 4) and arachnoid villi when CSFP and fluid retention are markedly elevated. A model is presented that links CSF and ISF homeostasis to coordinated fluxes of water and solutes at both the blood-CSF and blood-brain transport interfaces

Physiology of Blood–Brain Interfaces in Relation to Brain Disposition of Small Compounds and Macromolecules

The brain develops and functions within a strictly controlled environment resulting from the coordinated action of different cellular interfaces located between the blood and the extracellular fluids of the brain, which include the interstitial fluid and the cerebrospinal fluid (CSF). As a correlate, the delivery of pharmacologically active molecules and especially macromolecules to the brain is challenged by the barrier properties of these interfaces. Blood–brain interfaces comprise both the blood–brain barrier located at the endothelium of the brain microvessels and the blood–CSF barrier located at the epithelium of the choroid plexuses. Although both barriers develop extensive surface areas of exchange between the blood and the neuropil or the CSF, the molecular fluxes across these interfaces are tightly regulated. Cerebral microvessels acquire a barrier phenotype early during cerebral vasculogenesis under the influence of the Wnt/β-catenin pathway, and of recruited pericytes. Later in development, astrocytes also play a role in blood–brain barrier maintenance. The tight choroid plexus epithelium develops very early during embryogenesis. It is specified by various signaling molecules from the embryonic dorsal midline, such as bone morphogenic proteins, and grows under the influence of Sonic hedgehog protein. Tight junctions at each barrier comprise a distinctive set of claudins from the pore-forming and tightening categories that determine their respective paracellular barrier characteristics. Vesicular traffic is limited in the cerebral endothelium and abundant in the choroidal epithelium, yet without evidence of active fluid phase transcytosis. Inorganic ion transport is highly regulated across the barriers. Small organic compounds such as nutrients, micronutrients and hormones are transported into the brain by specific solute carriers. Other bioactive metabolites, lipophilic toxic xenobiotics or pharmacological agents are restrained from accumulating in the brain by several ATP-binding cassette efflux transporters, multispecific solute carriers, and detoxifying enzymes. These various molecular effectors differently distribute between the two barriers. Receptor-mediated endocytotic and transcytotic mechanisms are active in the barriers. They enable brain penetration of selected polypeptides and proteins, or inversely macromolecule efflux as it is the case for immnoglobulins G. An additional mechanism specific to the BCSFB mediates the transport of selected plasma proteins from blood into CSF in the developing brain. All these mechanisms could be explored and manipulated to improve macromolecule delivery to the brain

Biological roles of trace elements in the brain with special focus on Zn and Fe

International audienceINTRODUCTION: Many metals like iron (Fe), copper (Cu) or zinc (Zn) fulfil various essential biological functions and are thus vital for all living organisms. For instance, they play important roles in nervous tissue, participating in a wide range of processes such as neurotransmitter synthesis, myelination or synaptic transmission. STATE OF THE ART: As in other tissues, brain cells tightly control the concentration of metals but any excess or deficit can lead to deleterious responses and alter cognitive functions. Of note, certain metals such as Zn, Fe or Cu accumulate in specific brain structures over lifespan and several neurodegenerative diseases are associated with a dysregulation of the homeostatic mechanisms controlling the concentration of these cations. CONCLUSION AND PERSPECTIVES: This review will address some of the cellular and molecular processes controlling the entry and distribution of selected metals (mainly Zn and Fe) in the brain, as well as their roles in synaptic transmission, in the pathogenesis of some neurologic diseases such as Parkinson's disease and Alzheimer's disease, and their impact on cognitive functions