Mauro Ferrari

Shaping nano-/micro-particles for enhanced vascular interaction in laminar flows

Non-spherical nano-/micro-particles can drift laterally (hydrodynamic margination) in a linear laminar flow under the concurrent effect of hydrodynamic and inertial forces. Such a feature can be exploited in the rational design of particle-based intravascular and pulmonary delivery systems and for designing new flow fractioning systems for high-throughput particle separation. A general approach is presented to predict the marginating behavior of non-spherical particles. The lateral drift velocity is shown to depend on the particle Stokes number St(a) and to grow with the size, density and rotational inertia of the particle. Elongated particles, in particular, low aspect ratio discoidal particles, exhibit the largest propensity to marginate in a linear laminar flow. In the blood microcirculation, at low shear rates (S<100 s(-1)), non-spherical particles oscillate around their trajectory and margination can only be achieved through the application of external force fields (gravitational, magnetic); whereas for larger S (100 s(-1)<S<10(4) s(-1)), micrometer particles can achieve drift velocities in the order of 1-10 microm s(-1). In the pulmonary circulation, hydrodynamic margination can be observed even for sub-micrometer particles. Finally, the inherent propensity of non-spherical particles to drift laterally can be effectively exploited for designing microfluidic devices, based on the flow fractioning approach, for particle separation without using external lateral force fields.

Nanochannel technology for constant delivery of chemotherapeutics: beyond metronomic administration.

ABSTRACTPurposeThe purpose of this study is to demonstrate the long-term, controlled, zero-order release of low- and high-molecular weight chemotherapeutics through nanochannel membranes by exploiting the molecule-to-surface interactions presented by nanoconfinement.MethodsSilicon membranes were produced with nanochannels of 5, 13 and 20xa0nm using standardized industrial microfabrication techniques. The study of the diffusion kinetics of interferonα-2b and leuprolide was performed by employing UV diffusion chambers. The released amount in the sink reservoir was monitored by UV absorbance.ResultsContinuous zero-order release was demonstrated for interferonα-2b and leuprolide at release rates of 20 and 100xa0μg/day, respectively. The release rates exhibited by these membranes were verified to be in ranges suitable for human therapeutic applications.ConclusionsOur membranes potentially represent a viable nanotechnological approach for the controlled administration of chemotherapeutics intended to improve the therapeutic efficacy of treatment and reduce many of the side effects associated with conventional drug administration.

Probing the mechanical properties of TNF-α stimulated endothelial cell with atomic force microscopy.

TNF-α (tumor necrosis factor-α) is a potent pro-inflammatory cytokine that regulates the permeability of blood and lymphatic vessels. The plasma concentration of TNF-α is elevated (> 1 pg/mL) in several pathologies, including rheumatoid arthritis, atherosclerosis, cancer, pre-eclampsia; in obese individuals; and in trauma patients. To test whether circulating TNF-α could induce similar alterations in different districts along the vascular system, three endothelial cell lines, namely HUVEC, HPMEC, and HCAEC, were characterized in terms of 1) mechanical properties, employing atomic force microscopy; 2) cytoskeletal organization, through fluorescence microscopy; and 3) membrane overexpression of adhesion molecules, employing ELISA and immunostaining. Upon stimulation with TNF-α (10 ng/mL for 20 h), for all three endothelial cells, the mechanical stiffness increased by about 50% with a mean apparent elastic modulus of E ~5 ± 0.5 kPa (~3.3 ± 0.35 kPa for the control cells); the density of F-actin filaments increased in the apical and median planes; and the ICAM-1 receptors were overexpressed compared with controls. Collectively, these results demonstrate that sufficiently high levels of circulating TNF-α have similar effects on different endothelial districts, and provide additional information for unraveling the possible correlations between circulating pro-inflammatory cytokines and systemic vascular dysfunction.

Size of the nanovectors determines the transplacental passage in pregnancy: study in rats

OBJECTIVEnThe objective of the study was to examine whether the size of silicon nanovectors (SNVs) inhibits their entrance into the fetal circulation.nnnSTUDY DESIGNnPregnant rats were intravenously administered with SNVs or saline. The SNVs were spherical particles with 3 escalating diameters: 519 nm, 834 nm, and 1000 nm. The maternal and fetal distribution of SNVs was assessed.nnnRESULTSnIn animals that received 1000 or 834 nm SNV, silicon (Si) levels were significantly higher in the maternal organs vs the saline group, whereas the silicon levels in fetal tissues were similar to controls. However, in animals receiving 519 nm SNVs, fetal silicon levels were significantly higher in the SNV group compared with the saline group (5.93 ± 0.67 μg Si per organ vs 4.80 ± 0.33, P = .01).nnnCONCLUSIONnLarger SNVs do not cross the placenta to the fetus and, remaining within the maternal circulation, can serve as carriers for harmful medications in order to prevent fetal exposure.

Enhanced microcontact printing of proteins on nanoporous silica surface

We demonstrate porous silica surface modification, combined with microcontact printing, as an effective method for enhanced protein patterning and adsorption on arbitrary surfaces. Compared to conventional chemical treatments, this approach offers scalability and long-term device stability without requiring complex chemical activation. Two chemical surface treatments using functionalization with the commonly used 3-aminopropyltriethoxysilane (APTES) and glutaraldehyde (GA) were compared with the nanoporous silica surface on the basis of protein adsorption. The deposited thickness and uniformity of porous silica films were evaluated for fluorescein isothiocyanate (FITC)-labeled rabbit immunoglobulin G (R-IgG) protein printed onto the substrates via patterned polydimethlysiloxane (PDMS) stamps. A more complete transfer of proteins was observed on porous silica substrates compared to chemically functionalized substrates. A comparison of different pore sizes (4-6 nm) and porous silica thicknesses (96-200 nm) indicates that porous silica with 4 nm diameter, 57% porosity and a thickness of 96 nm provided a suitable environment for complete transfer of R-IgG proteins. Both fluorescence microscopy and atomic force microscopy (AFM) were used for protein layer characterizations. A porous silica layer is biocompatible, providing a favorable transfer medium with minimal damage to the proteins. A patterned immunoassay microchip was developed to demonstrate the retained protein function after printing on nanoporous surfaces, which enables printable and robust immunoassay detection for point-of-care applications.

Device for rapid and agile measurement of diffusivity in micro- and nanochannels.

The lack of a viable theory for describing diffusivity when fluids are confined at the micro- and nanoscale [Ladero et al. Chem. Eng. Sci.2007, 62, 666-678; Deen AIChE J.1987, 33, 1409-1425] has necessitated accurate measurement of diffusivity (D) [Jin and Chen Chromatographia2000, 52, 17-21; Nie et al. Science1994, 266, 1018-1021; Durand et al. Anal. Chem.2009, 81, 5407-5412], crucial for a host of micro- and nanofluidic technologies [Grattoni et al. Curr. Pharm. Biotechnol.2010, 11, 343-365]. We demonstrate a rapid and agile method for the direct measurement of diffusivity in a system possessing 10(4) to 10(5) precisely fabricated channels with characteristic sizes (β) ranging from micro- to nanometers. Custom chambers allowed us to measure the diffusivity in a closed unperturbed system using UV/vis spectroscopy. D was measured for rhodamine B (RhoB) in aqueous solution in channels of 200 and 1 μm, as well as 13 and 5.7 nm. The observed logarithmic scaling of diffusivity with β, in close agreement with prior experiments, but far from theoretical prediction, surprisingly highlights that diffusivity is significantly altered even at the microscale. Accurate measurement of D by reducing the size of the source reservoir by 3 orders of magnitude (from 150 μL to 910 nL) proves that a substantial reduction in measurement time (from 7 days to 40 min) can be achieved. Our design thus is ready for rapid translation into a standard analytical tool--useful for multiple applications.

Application of physicochemically modified silicon substrates as reverse phase protein microarrays

Physicochemically modified silicon substrates can provide a high quality alternative to nitrocellulose-coated glass slides for use in reverse-phase protein microarrays. Enhancement of protein microarray sensitivities is an important goal, especially because molecular targets within patient tissues exist in low abundance. The ideal array substrate has a high protein binding affinity and low intrinsic background signal. Silicon, which has low intrinsic autofluorescence, is being explored as a potential microarray surface. In a previous paper ( Nijdam , A. J. ; Cheng , M. M.-C. ; Fedele , R. ; Geho , D. H. ; Herrmann , P. ; Killian , K. ; Espina , V. ; Petricoin , E. F. ; Liotta , L. A. ; Ferrari , M. Physicochemically Modified Silicon as Substrate for Protein Microarrays . Biomaterials 2007 , 28 , 550 - 558 ), it is shown that physicochemical modification of silicon substrates increases the binding of protein to silicon to a level comparable with that of nitrocellulose. Here, we apply such substrates in a reverse-phase protein microarray setting in two model systems.

Vectoring siRNA therapeutics into the clinic.

Nanoparticles are a promising vector-based strategy for the therapeutic administration of small interfering (si)RNA because they protect siRNA from nuclease degradation. A recent phase I study employed transferrin–targeted nanovectors to demonstrate RNA interference mechanisms in a melanoma patient. Multifunctional nanoparticles are providing patient-specific biodistributions of systemically administered siRNA.