Paulo Henrique Jacob Silva
EPFL STI IMX SUNMIL
MXG 034 (Bâtiment MXG)
Station 12
1015 Lausanne
Web site: Site web: https://sunmil.epfl.ch
Formation
Bachelor of Science
Materials Science and Engineering
MIT
2006
PhD
Materials Science
EPFL
2016
Publications
Sélection de publications
| Urszula Cendrowska*, Paulo Jacob Silva*, Nadine Ait-Bouziad, Marie Müller, Zekiye Pelin Guven, Sophie Vieweg, Anass Chiki, Lynn Radamaker, Senthil T. Kumar, Marcus Fändrich, Francesco Tavanti, Maria Cristina Menziani, Alfredo Alexander-Katz, View ORCID ProfileFrancesco Stellacci, and Hilal A. Lashuel
The ability of proteins to self-assemble into different types of fibrils with distinct morphologies has been linked to the pathological and clinical heterogeneity of amyloid diseases such as Alzheimer’s disease and Parkinson’s disease. Here, we describe nanoparticles (NPs) that efficiently label amyloid fibrils produced in vitro or isolated from postmortem tissues, under hydrating conditions and in such a way as to unmask their polymorphism and morphological features. Using these NPs, we show that pathological aggregates exhibit exceptional morphological homogeneity compared with amyloid fibrils produced in vitro, consistent with the emerging view that the physiologic milieu is a key determinant of amyloid fibril strains. These advances pave the way for elucidating the structural basis of amyloid strains and toxicity. |
Unraveling the complexity of amyloid polymorphism using gold nanoparticles and cryo-EM |
| Valeria Cagno, Patrizia Andreozzi, Marco D’Alicarnasso, Paulo Jacob Silva, Marie Mueller, Marie Galloux, Ronan Le Goffic, Samuel T. Jones, Marta Vallino, Jan Hodek, Jan Weber, Soumyo Sen, Emma-Rose Janeček, Ahmet Bekdemir, Barbara Sanavio, Chiara Martinelli, Manuela Donalisio, Marie-Anne Rameix Welti, Jean-Francois Eleouet, Yanxiao Han, Laurent Kaiser, Lela Vukovic, Caroline Tapparel, Petr Král, Silke Krol, David Lembo & Francesco Stellacci Viral infections kill millions yearly. Available antiviral drugs are virus-specific and active against a limited panel of human pathogens. There are broad-spectrum substances that prevent the first step of virus–cell interaction by mimicking heparan sulfate proteoglycans (HSPG), the highly conserved target of viral attachment ligands (VALs). The reversible binding mechanism prevents their use as a drug, because, upon dilution, the inhibition is lost. Known VALs are made of closely packed repeating units, but the aforementioned substances are able to bind only a few of them. We designed antiviral nanoparticles with long and flexible linkers mimicking HSPG, allowing for effective viral association with a binding that we simulate to be strong and multivalent to the VAL repeating units, generating forces (∼190 pN) that eventually lead to irreversible viral deformation. Virucidal assays, electron microscopy images, and molecular dynamics simulations support the proposed mechanism. These particles show no cytotoxicity, and in vitro nanomolar irreversible activity against herpes simplex virus (HSV), human papilloma virus, respiratory syncytial virus (RSV), dengue and lenti virus. They are active ex vivo in human cervicovaginal histocultures infected by HSV-2 and in vivo in mice infected with RSV. |
Broad-spectrum non-toxic antiviral nanoparticles with a virucidal inhibition mechanism |
| Zekiye P. Guven*, Paulo H. Jacob Silva*, Zhi Luo, Urszula B. Cendrowska, Matteo Gasbarri, Samuel T. Jones, Francesco Stellacci Gold nanoparticles covered with a mixture of 1-octanethiol (OT) and 11-mercapto-1-undecane sulfonic acid (MUS) have been extensively studied because of their interactions with cell membranes, lipid bilayers, and viruses. The hydrophilic ligands make these particles colloidally stable in aqueous solutions and the combination with hydrophobic ligands creates an amphiphilic particle that can be loaded with hydrophobic drugs, fuse with the lipid membranes, and resist nonspecific protein adsorption. Many of these properties depend on nanoparticle size and the composition of the ligand shell. It is, therefore, crucial to have a reproducible synthetic method and reliable characterization techniques that allow the determination of nanoparticle properties and the ligand shell composition. Here, a one-phase chemical reduction, followed by a thorough purification to synthesize these nanoparticles with diameters below 5 nm, is presented. The ratio between the two ligands on the surface of the nanoparticle can be tuned through their stoichiometric ratio used during synthesis. We demonstrate how various routine techniques, such as transmission electron microscopy (TEM), nuclear magnetic resonance (NMR), thermogravimetric analysis (TGA), and ultraviolet-visible (UV-Vis) spectrometry, are combined to comprehensively characterize the physicochemical parameters of the nanoparticles. |
Synthesis and Characterization of Amphiphilic Gold Nanoparticles |
| Maria Pelliccia, Patrizia Andreozzi, Jayson Paulose, Marco D’Alicarnasso, Valeria Cagno, Manuela Donalisio, Andrea Civra, Rebecca M. Broeckel, Nicole Haese, Paulo Jacob Silva, Randy P. Carney, Varpu Marjomäki, Daniel N. Streblow, David Lembo, Francesco Stellacci, Vincenzo Vitelli & Silke Krol Up to 80% of the cost of vaccination programmes is due to the cold chain problem (that is, keeping vaccines cold). Inexpensive, biocompatible additives to slow down the degradation of virus particles would address the problem. Here we propose and characterize additives that, already at very low concentrations, improve the storage time of adenovirus type 5. Anionic gold nanoparticles (10−8–10−6 M) or polyethylene glycol (PEG, molecular weight ∼8,000 Da, 10−7–10−4 M) increase the half-life of a green fluorescent protein expressing adenovirus from ∼48 h to 21 days at 37 °C (from 7 to >30 days at room temperature). They replicate the known stabilizing effect of sucrose, but at several orders of magnitude lower concentrations. PEG and sucrose maintained immunogenicity in vivo for viruses stored for 10 days at 37 °C. To achieve rational design of viral-vaccine stabilizers, our approach is aided by simplified quantitative models based on a single rate-limiting step. |
Additives for vaccine storage to improve thermal stability of adenoviruses from hours to months |
| Reid C. Van Lehn, Maria Ricci, Paulo H.J. Silva, Patrizia Andreozzi, Javier Reguera, Kislon Voïtchovsky, Francesco Stellacci & Alfredo Alexander-Katz Recent work has demonstrated that charged gold nanoparticles (AuNPs) protected by an amphiphilic organic monolayer can spontaneously insert into the core of lipid bilayers to minimize the exposure of hydrophobic surface area to water. However, the kinetic pathway to reach the thermodynamically stable transmembrane configuration is unknown. Here, we use unbiased atomistic simulations to show the pathway by which AuNPs spontaneously insert into bilayers and confirm the results experimentally on supported lipid bilayers. The critical step during this process is hydrophobic–hydrophobic contact between the core of the bilayer and the monolayer of the AuNP that requires the stochastic protrusion of an aliphatic lipid tail into solution. This last phenomenon is enhanced in the presence of high bilayer curvature and closely resembles the putative pre-stalk transition state for vesicle fusion. To the best of our knowledge, this work provides the first demonstration of vesicle fusion-like behaviour in an amphiphilic nanoparticle system. |
Lipid tail protrusions mediate the insertion of nanoparticles into model cell membranes |
| Randy P. Carney, Yann Astier, Tamara M. Carney, Kislon Voïtchovsky, Paulo H. Jacob Silva, and Francesco Stellacci Understanding as well as rapidly screening the interaction of nanoparticles with cell membranes is of central importance for biological applications such as drug and gene delivery. Recently, we have shown that “striped” mixed-monolayer-coated gold nanoparticles spontaneously penetrate a variety of cell membranes through a passive pathway. Here, we report an electrical approach to screen and readily quantify the interaction between nanoparticles and bilayer lipid membranes. Membrane adsorption is monitored through the capacitive increase of suspended planar lipid membranes upon fusion with nanoparticles. We adopt a Langmuir isotherm model to characterize the adsorption of nanoparticles by bilayer lipid membranes and extract the partition coefficient, K, and the standard free energy gain by this spontaneous process, for a variety of sizes of cell-membrane-penetrating nanoparticles. We believe that the method presented here will be a useful qualitative and quantitative tool to determine nanoparticle interaction with lipid bilayers and consequently with cell membranes. |
Electrical Method to Quantify Nanoparticle Interaction with Lipid Bilayers |
| Fernanda Sousa, Barbara Sanavio, Alessandra Saccani, Yun Tang§∥, Ileana Zucca, Tamara M. Carney, Alfonso Mastropietro, Paulo H. Jacob Silva, Randy P. Carney, Kurt Schenk, Arash O. Omrani, Ping Huang∇, Lin Yang, Henrik M. Rønnow, Francesco Stellacci, and Silke Krol Nanoparticle-based magnetic resonance imaging T2 negative agents are of great interest, and much effort is devoted to increasing cell-loading capability while maintaining low cytotoxicity. Herein, two classes of mixed-ligand protected magnetic-responsive, bimetallic gold/iron nanoparticles (Au/Fe NPs) synthesized by a two-step method are presented. Their structure, surface composition, and magnetic properties are characterized. The two classes of sulfonated Au/Fe NPs, with an average diameter of 4 nm, have an average atomic ratio of Au to Fe equal to 7 or 8, which enables the Au/Fe NPs to be superparamagnetic with a blocking temperature of 56 K and 96 K. Furthermore, preliminary cellular studies reveal that both Au/Fe NPs show very limited toxicity. MRI phantom experiments show that r2/r1 ratio of Au/Fe NPs is as high as 670, leading to a 66% reduction in T2 relaxation time. These nanoparticles provide great versatility and potential for nanoparticle-based diagnostics and therapeutic applications and as imaging contrast agents. |
Superparamagnetic Nanoparticles as High Efficiency Magnetic Resonance Imaging T2 Contrast Agent |
| Quy Khac Ong, Javier Reguera, Paulo Jacob Silva, Mauro Moglianetti, Kellen Harkness, Maria Longobardi, Kunal S. Mali, Christoph Renner, Steven De Feyter, and Francesco Stellacci Gold nanoparticles protected by a binary mixture of thiolate molecules have a ligand shell that can spontaneously separate into nanoscale domains. Complex morphologies arise in such ligand shells, including striped, patchy, and Janus domains. Characterization of these morphologies remains a challenge. Scanning tunneling microscopy (STM) imaging has been one of the key approaches to determine these structures, yet the imaging of nanoparticles’ surfaces faces difficulty stemming from steep surface curvature, complex molecular structures, and the possibility of imaging artifacts in the same size range. Images obtained to date have lacked molecular resolution, and only domains have been resolved. There is a clear need for images that resolve the molecular arrangement that leads to domain formation on the ligand shell of these particles. Herein we report an advance in the STM imaging of gold nanoparticles, revealing some of the molecules that constitute the domains in striped and Janus gold nanoparticles. We analyze the images to determine molecular arrangements on parts of the particles, highlight molecular “defects” present in the ligand shell, show persistence of the features across subsequent images, and observe the transition from quasi-molecular to domain resolution. The ability to resolve single molecules in the ligand shell of nanoparticles could lead to a more comprehensive understanding of the role of the ligand structure in determining the properties of mixed-monolayer-protected gold nanoparticles. |
High-Resolution Scanning Tunneling Microscopy Characterization of Mixed Monolayer Protected Gold Nanoparticles |
| rabhani U. Atukorale, Yu-Sang Yang, Ahmet Bekdemir, Randy P. Carney, Paulo J. Silva, Nicki Watson, Francesco Stellacci and Darrell J. Irvine Erythrocytes are attractive as potential cell-based drug carriers because of their abundance and long lifespan in vivo. Existing methods for loading drug cargos into erythrocytes include hypotonic treatments, electroporation, and covalent attachment onto the membrane, all of which require ex vivo manipulation. Here, we characterized the properties of amphiphilic gold nanoparticles (amph-AuNPs), comprised of a ∼2.3 nm gold core and an amphiphilic ligand shell, which are able to embed spontaneously within erythrocyte membranes and might provide a means to load drugs into red blood cells (RBCs) directly in vivo. Particle interaction with RBC membranes occurred rapidly at physiological temperature. We further show that amph-AuNP uptake by RBCs was limited by the glycocalyx and was particularly influenced by sialic acids on cell surface proteoglycans. Using a reductionist model membrane system with synthetic lipid vesicles, we confirmed the importance of membrane fluidity and the glycocalyx in regulating amph-AuNP/membrane interactions. These results thus provide evidence for the interaction of amph-AuNPs with erythrocyte membranes and identify key membrane components that govern this interaction, providing a framework for the development of amph-AuNP-carrying erythrocyte ‘pharmacytes’ in vivo. |
Influence of the glycocalyx and plasma membrane composition on amphiphilic gold nanoparticle association with erythrocytes |
| Alysia Cox, Patrizia Andreozzi, Roberta Dal Magro, Fabio Fiordaliso, Alessandro Corbelli, Laura Talamini, Clizia Chinello, Francesca Raimondo, Fulvio Magni, Maria Tringali, Silke Krol, Paulo Jacob Silva, Francesco Stellacci, Massimo Masserini, and Francesca Re Engineered nanoparticles offer the chance to improve drug transport and delivery through biological barriers, exploiting the possibility to leave the blood circulation and traverse the endothelial vascular bed, blood–brain barrier (BBB) included, to reach their target. It is known that nanoparticles gather molecules on their surface upon contact with biological fluids, forming the “protein corona”, which can affect their fate and therapeutic/diagnostic performance, yet no information on the corona’s evolution across the barrier has been gathered so far. Using a cellular model of the BBB and gold nanoparticles, we show that the composition of the corona undergoes dramatic quantitative and qualitative molecular modifications during passage from the “blood” to the “brain” side, while it is stable once beyond the BBB. Thus, we demonstrate that the nanoparticle corona dynamically and drastically evolves upon crossing the BBB and that its initial composition is not predictive of nanoparticle fate and performance once beyond the barrier at the target organ. |
Evolution of Nanoparticle Protein Corona across the Blood–Brain Barrier |