Sandra Rocha

Gut Microbiota Regulate Motor Deficits and Neuroinflammation in a Model of Parkinson’s Disease

The intestinal microbiota influence neurodevelopment, modulate behavior, and contribute to neurological disorders. However, a functional link between gut bacteria and neurodegenerative diseases remains unexplored. Synucleinopathies are characterized by aggregation of the protein α-synuclein (αSyn), often resulting in motor dysfunction as exemplified by Parkinsons disease (PD). Using mice that overexpress αSyn, we report herein that gut microbiota are required for motor deficits, microglia activation, and αSyn pathology. Antibiotic treatment ameliorates, while microbial re-colonization promotes, pathophysiology in adult animals, suggesting that postnatal signaling between the gut and the brain modulates disease. Indeed, oral administration of specific microbial metabolites to germ-free mice promotes neuroinflammation and motor symptoms. Remarkably, colonization of αSyn-overexpressing mice with microbiota from PD-affected patients enhances physical impairments compared to microbiota transplants from healthy human donors. These findings reveal that gut bacteria regulate movement disorders in mice and suggest that alterations in the human microbiome represent a risk factor for PD.

Influence of fluorinated and hydrogenated nanoparticles on the structure and fibrillogenesis of amyloid beta-peptide.

Peptide aggregation in amyloid fibrils is implicated in the pathogenesis of several diseases such as Alzheimers disease. There is a strong correlation between amyloid fibril formation and a decrease in conformational stability of the native state. Amyloid-beta peptide (Abeta), the aggregating peptide in Alzheimers disease, is natively unfolded. The deposits found in Alzheimers disease are composed of Abeta fibrillar aggregates rich in beta-sheet structure. The influence of fluorinated complexes on the secondary structure and fibrillogenesis of Abeta peptide was studied by circular dichroism (CD) spectroscopy and transmission electron microscopy (TEM). CD spectra show that complexes of polyampholyte and fluorinated dodecanoic acid induce alpha-helix structure in Abeta, but their hydrogenated analogous lead to beta-sheet formation and aggregation. The fluorinated nanoparticles with highly negative zeta potential and hydrophobic fluorinated core have the fundamental characteristics to prevent Abeta fibrillogenesis.

Epigallocatechin gallate-loaded polysaccharide nanoparticles for prostate cancer chemoprevention

AIMS Polysaccharide nanoparticles were studied as drug delivery vehicles for chemopreventive agents. MATERIALS & METHODS Green tea polyphenol epigallocatechin-3-gallate (EGCG) was incorporated into a carbohydrate matrix of gum arabic and maltodextrin with an encapsulation efficiency of approximately 85%. RESULTS Encapsulated EGCG retained its biological activity, reducing the cell viability and inducing apoptosis of Du145 prostate cancer cells. Clonogenic assay demonstrated that encapsulation of EGCG enhanced its inhibitory effect on cell proliferation (10-20%) at lower concentrations (1-2 µM), compared with free EGCG. CONCLUSION This study highlights the use of polysaccharide nanoparticles in chemoprevention as they can be used to deliver natural antioxidants capable of inhibiting steps of the tumorigenesis process.

Adsorption and Diffusion of Plasma Proteins on Hydrophilic and Hydrophobic Surfaces: Effect of Trifluoroethanol on Protein Structure

The aim of this work was to investigate the conformational changes and diffusion of adsorbed proteins (immunoglobulin G (IgG), fibrinogen (Fib) and human serum albumin (HSA)) on hydrophilic quartz and hydrophobized quartz (octadecyltrichlorosilane (OTS)) surfaces. Circular dichroism spectroscopy measurements have shown that IgG is the most stable protein after adsorption on hydrophilic quartz, whereas HSA and Fib unfold. The structural changes are dependent on adsorption time, initial protein concentration in bulk, and surface chemistry. The effect of trifluoroethanol (TFE) in recovering the original protein structure after adsorption was analyzed by total internal reflection fluorescence and fluorescence recovery after photobleaching (TIRF-FRAP). TIRF-FRAP experiments revealed a strong dependence of the surface chemistry on protein diffusion coefficients: proteins diffuse 4 times slower on hydrophobic surfaces than on hydrophilic surfaces. The diffusion coefficient of TFE at hydrophobic surfaces is 2 orders magnitude higher than at hydrophilic surfaces. However, protein desorption occurs faster on hydrophilic quartz than on OTS, proving that the strength of protein-surface interaction is weaker at hydrophilic surfaces. This result shows that desorption is determined by surface/protein chemistry and not by mass transfer limitations. FTIR-ATR results demonstrated that TFE interaction with adsorbed proteins is stronger at hydrophilic surfaces than at hydrophobic surfaces.

Targeting nanoparticles across the blood-brain barrier with monoclonal antibodies

Development of therapeutics for brain disorders is one of the more difficult challenges to be overcome by the scientific community due to the inability of most molecules to cross the blood-brain barrier (BBB). Antibody-conjugated nanoparticles are drug carriers that can be used to target encapsulated drugs to the brain endothelial cells and have proven to be very promising. They significantly improve the accumulation of the drug in pathological sites and decrease the undesirable side effect of drugs in healthy tissues. We review the systems that have demonstrated promising results in crossing the BBB through receptor-mediated endocytic mechanisms for the treatment of neurodegenerative disorders such as Alzheimers and Parkinsons disease.

Lipid/particle assemblies based on maltodextrin-gum arabic core as bio-carriers.

A novel system to carry and protect epigallocatechin gallate (EGCG), an antioxidant from the green tea, is reported. The system consists of maltodextrin and gum arabic nanoparticles coated with egg-yolk l-alpha-phosphatidylcholine (Egg-PC)/stearylamine (SA) bilayers. In this study, the polysaccharide core was produced by homogenization followed by spray-drying. The lipid coating was performed by the lipid film hydration method. The polysaccharide core revealed negative zeta potential, which changed to opposite signs after lipid coating. The presence of lipid layers was evidenced by cryogenic-transmission (cryo-TEM) and scanning (cryo-SEM) electron microscopy studies. An increase in size was observed after lipid coating as determined by dynamic light scattering (DLS). Atomic force microscopy (AFM) demonstrated that the polysaccharide core provides high resistance to mechanical strength. The lipid/particle assemblies show high retention efficiency of EGCG at physiological pH, opening the possibility of their use for delivery and controlled release of tea catechins.

Design and biological activity of β-sheet breaker peptide conjugates

The sequence LPFFD (iAbeta(5)) prevents amyloid-beta peptide (Abeta) fibrillogenesis and neurotoxicity, hallmarks of Alzheimers disease (AD), as previously demonstrated. In this study iAbeta(5) was covalently linked to poly(ethylene glycol) (PEG) and the activity of conjugates was assessed and compared to the activity of the peptide alone by in vitro studies. The conjugates were characterized by MALDI-TOF. Competition binding assays established that conjugates retained the ability to bind Abeta with similar strength as iAbeta(5). Transmission electron microscopy analysis showed that iAbeta(5) conjugates inhibited amyloid fibril formation, which is in agreement with binding properties observed for the conjugates towards Abeta. The conjugates were also able to prevent amyloid-induced cell death, as evaluated by activation of caspase 3. These results demonstrated that the biological activity of iAbeta(5) is not affected by the pegylation process.

Carbon-core silver-shell nanodots as sensitizers for phototherapy and radiotherapy

Spherical carbon nanoparticles (carbon nanodots) with a silver shell were investigated as potential sensitizing agents. The cytotoxicity of the combination of ultraviolet radiation or x-rays with the nanodots was examined in cancer cells in vitro. The cell viability decreased following the exposure to the radiation. The carbon nanodots enhanced the radiation effects by significantly reducing the amount of surviving cells compared to that of the cells exposed only to the radiation. Carbon-core silver-shell nanodots can be proposed as a bimodal sensitization platform for biological and medicinal applications employing non-ionizing or ionizing radiation.

Nanostructure of polysaccharide complexes.

The interaction of gum arabic (GA) with chitosan (Ch) of different degree of deacetylation was studied by turbidity measurements, dynamic light scattering and atomic force microscopy. The structure of the complexes was found to be directly related to the charge density of chitosan molecules. Gum arabic and chitosan with a degree of deacetylation of 75% form soluble complexes with a loosely globular structure of about 250 nm, at weight ratios up to 1.2, if the concentrations are kept low (total biopolymer concentration up to 0.06%). If chitosan has a higher charge density (degree of deacetylation of 93%), colloidal particles are formed, independently of the polymer concentration or ratio. At low concentrations and GA/Ch ratios of 1 or 1.2, the particles have diameters of 200-250 nm. The formation of soluble complexes is attributed to a chitosan lower charge density and the presence of non-charged monomers, which prevent the efficient self-assembly of the macromolecules.

Cellular uptake of PLGA nanoparticles targeted with anti-amyloid and anti-transferrin receptor antibodies for Alzheimer’s disease treatment

During the last few decades, relevant efforts have been reported to design nanocarriers for drug transport through the blood brain barrier (BBB). New drugs, such as peptide iAβ5, capable to inhibit the aggregates associated with Alzheimeŕs disease (AD) are being tested but the most frequent drawback is to reach the brain in the desired concentrations due to the low BBB permeability-surface area. Our approach, as a proof of concept to improve drug transport through the BBB, is based on poly(lactic-co-glycolic acid) (PLGA) nanoparticles with surface functionalized with anti-transferrin receptor monoclonal antibody (OX26) and anti-Aβ (DE2B4) to deliver encapsulated iAβ5 into the brain. Porcine brain capillary endothelial cells (PBCECs) were used as a BBB model to evaluate the system efficacy and toxicity. The uptake of immune nanoparticles with a controlled delivery of the peptide iAβ5 was substantially increased compared to the nanoparticles (NPs) without monoclonal antibody functionalization.