Virginia Pensabene

Recreating blood-brain barrier physiology and structure on chip: A novel neurovascular microfluidic bioreactor.

The blood-brain barrier (BBB) is a critical structure that serves as the gatekeeper between the central nervous system and the rest of the body. It is the responsibility of the BBB to facilitate the entry of required nutrients into the brain and to exclude potentially harmful compounds; however, this complex structure has remained difficult to model faithfully in vitro. Accurate in vitro models are necessary for understanding how the BBB forms and functions, as well as for evaluating drug and toxin penetration across the barrier. Many previous models have failed to support all the cell types involved in the BBB formation and/or lacked the flow-created shear forces needed for mature tight junction formation. To address these issues and to help establish a more faithful in vitro model of the BBB, we have designed and fabricated a microfluidic device that is comprised of both a vascular chamber and a brain chamber separated by a porous membrane. This design allows for cell-to-cell communication between endothelial cells, astrocytes, and pericytes and independent perfusion of both compartments separated by the membrane. This NeuroVascular Unit (NVU) represents approximately one-millionth of the human brain, and hence, has sufficient cell mass to support a breadth of analytical measurements. The NVU has been validated with both fluorescein isothiocyanate (FITC)-dextran diffusion and transendothelial electrical resistance. The NVU has enabled in vitro modeling of the BBB using all human cell types and sampling effluent from both sides of the barrier.

Neurovascular unit on a chip: implications for translational applications

The blood-brain barrier (BBB) dynamically controls exchange between the brain and the body, but this interaction cannot be studied directly in the intact human brain or sufficiently represented by animal models. Most existing in vitro BBB models do not include neurons and glia with other BBB elements and do not adequately predict drug efficacy and toxicity. Under the National Institutes of Health Microtissue Initiative, we are developing a three-dimensional, multicompartment, organotypic microphysiological system representative of a neurovascular unit of the brain. The neurovascular unit system will serve as a model to study interactions between the central nervous system neurons and the cerebral spinal fluid (CSF) compartment, all coupled to a realistic blood-surrogate supply and venous return system that also incorporates circulating immune cells and the choroid plexus. Hence all three critical brain barriers will be recapitulated: blood-brain, brain-CSF, and blood-CSF. Primary and stem cell-derived human cells will interact with a variety of agents to produce critical chemical communications across the BBB and between brain regions. Cytomegalovirus, a common herpesvirus, will be used as an initial model of infections regulated by the BBB. This novel technological platform, which combines innovative microfluidics, cell culture, analytical instruments, bioinformatics, control theory, neuroscience, and drug discovery, will replicate chemical communication, molecular trafficking, and inflammation in the brain. The platform will enable targeted and clinically relevant nutritional and pharmacologic interventions for or prevention of such chronic diseases as obesity and acute injury such as stroke, and will uncover potential adverse effects of drugs. If successful, this project will produce clinically useful technologies and reveal new insights into how the brain receives, modifies, and is affected by drugs, other neurotropic agents, and diseases.

Evaluation of friction enhancement through soft polymer micro-patterns in active capsule endoscopy

Capsule endoscopy is an emerging field in medical technology. Despite very promising innovations, some critical issues are yet to be addressed, such as the management and possible exploitation of the friction in the gastrointestinal environment in order to control capsule locomotion more actively. This paper presents the fabrication and testing of bio-inspired polymeric micro-patterns, which are arrays of cylindrical pillars fabricated via soft lithography. The aim of the work is to develop structures that enhance the grip between an artificial device and the intestinal tissue, without injuring the mucosa. In fact, the patterns are intended to be mounted on microfabricated legs of a capsule robot that is able to move actively in the gastrointestinal tract, thus improving the robots traction ability. The effect of micro-patterned surfaces on the leg-slipping behaviour on colon walls was investigated by considering both different pillar dimensions and the influence of tissue morphology. Several in vitro tests on biological samples demonstrated that micro-patterns of pillars made from a soft polymer with an aspect ratio close to 1 enhanced friction by 41.7% with regard to flat surfaces. This work presents preliminary modelling of the friction and adhesion forces in the gastrointestinal environment and some design guidelines for endoscopic devices.

Free-Standing Poly(L-lactic acid) Nanofilms Loaded with Superparamagnetic Nanoparticles

Freely suspended nanocomposite thin films based on soft polymers and functional nanostructures have been widely investigated for their potential application as active elements in microdevices. However, most studies are focused on the preparation of nanofilms composed of polyelectrolytes and charged colloidal particles. Here, a new technique for the preparation of poly(l-lactic acid) free-standing nanofilms embeddidng superparamagnetic iron oxide nanoparticles is presented. The fabrication process, based on a spin-coating deposition approach, is described, and the influence of each production parameter on the morphology and magnetic properties of the final structure is investigated. Superparamagnetic free-standing nanofilms were obtained, as evidenced by a magnetization hysteresis measurement performed with a superconducting quantum interference device (SQUID). Nanofilm surface morphology and thickness were evaluated by atomic force microscopy (AFM), and the nanoparticle dispersion inside the composites was investigated by transmission electron microscopy (TEM). These nanofilms, composed of a biodegradable polyester and remotely controllable by external magnetic fields, are promising candidates for many potential applications in the biomedical field.

Dispersion of Multi‐walled Carbon Nanotubes in Aqueous Pluronic F127 Solutions for Biological Applications

Because mass‐produced carbon nanotubes (CNTs) are strongly aggregated and highly hydrophobic, processes to make them water soluble are required for biological applications. Suspensions in surfactant solutions are often employed. Among these, Pluronic F127 appear to be highly biocompatible if used at low concentrations. Starting from these results, this work involves a systematic study to clarify the dispersion behaviour of CNTs in Pluronic F127. The results suggest a two‐step process: first, the bundles disaggregate, kinetically driven by the energy supplied to the system; second, they disperse (surfactant adsorption), thermodynamically driven by the surfactant concentration. The dispersion reaction data are well fitted by a first‐order kinetics reaction. By performing a pretreatment step, consisting of stirring at 70°C, the achieved concentration of CNTs in solution is twice that of the traditional process. The proposed procedure provides an optimal compromise between a low Pluronic concentration and a high CNT concentration.

Carbon nanotube-enhanced cell electropermeabilisation.

The use of controlled electric fields to facilitate cell permeabilisation for enhanced cellular uptake of molecules is well established. The main limitation to the application of this technology in clinical practice is the requirements of high voltages which cause significant cell death in the target tissue. This paper presents a new modality for cell electro-permeabilisation based on the use of carbon nanotubes (CNTs) and external static electric fields. An explanation of the results based on the dielectric response of multiwall CNTs (MWCNTs) to these electric fields is proposed. The experimental data obtained indicate that this method of CNT-enhanced electro-permeabilisation provides an effective means of lowering the electric field voltage required for repairable cell electro-permeabilisation to below 50V/cm and with an efficiency exceeding 80%.

Cell Creeping and Controlled Migration by Magnetic Carbon Nanotubes

Carbon nanotubes (CNTs) are tubular nanostructures that exhibit magnetic properties due to the metal catalyst impurities entrapped at their extremities during fabrication. When mammalian cells are cultured in a CNT-containing medium, the nanotubes interact with the cells, as a result of which, on exposure to a magnetic field, they are able to move cells towards the magnetic source. In the present paper, we report on a model that describes the dynamics of this mammalian cell movement in a magnetic field consequent on CNT attachment. The model is based on Bell’s theory of unbinding dynamics of receptor-ligand bonds modified and validated by experimental data of the movement dynamics of mammalian cells cultured with nanotubes and exposed to a magnetic field, generated by a permanent magnet, in the vicinity of the cell culture wells. We demonstrate that when the applied magnetic force is below a critical value (about Fc ≈ 10−11 N), the cell ‘creeps’ very slowly on the culture dish at a very low velocity (10–20 nm/s) but becomes detached from the substrate when this critical magnetic force is exceeded and then move towards the magnetic source.

Design and development of a soft magnetically-propelled swimming microrobot

A novel approach for the design of magnetically-propelled microrobots is proposed as an effective solution for swimming in a liquid medium. While intrinsic neutral buoyancy of a microrobot per se simplifies propulsion in the liquid environments, softness makes it compliant with delicate environments, such as the human body, thus guaranteeing a safe interaction with soft structures. With this aim, two groups of soft microrobots with paramagnetic and ferromagnetic behaviors were designed, fabricated and their features were experimentally analyzed. In agreement with the theoretical predictions, in the performed trials the ferromagnetic microrobots showed orientation capabilities in response to the magnetic field that could not be achieved by the paramagnetic one. Moreover, it was observed that the ferromagnetic microrobot could reach higher speed values (maximum value of 0.73 body length/s) than the paramagnetic prototype.

Neuroblastoma Cells Displacement by Magnetic Carbon Nanotubes

In this paper, as-produced multiwall carbon nanotubes (MWNTs) have been analyzed by scanning electron microscopy and energy dispersive X-ray spectrometry, revealing the presence of Fe, Al, and Zn residuals and impurities. MWNTs have then been dispersed in Pluronic F127 aqueous solution and used to seed neuroblastoma cell lines (HN9.10e and SH-SY5Y) for three days. We found that MWNTs interact with cells and induce, under a permanent constant magnetic field, the cell displacement toward the magnetic source.

A pilot study on a new anchoring mechanism for surgical applications based on mucoadhesives

Abstract In order to minimize the invasiveness of laparoscopic surgery, different techniques are emerging from research to clinical practice. Whether the incision is performed on the outside – as in Single Port Laparoscopy (SPL) – or on the inside – as in Natural Orifice Transluminal Endoscopic Surgery (NOTES) – of the patients body, inserting and operating all the instruments from a single access site seems to be the next challenge in surgery. Magnetic guidance has been recently proposed for controlling surgical tools deployed from a single access. However, the exponential drop of magnetic field with distance makes this solution suitable only for the upper side of the abdominal cavity in nonobese patients. In the present paper we introduce a polymeric anchoring mechanism to lock surgical assistive tools inside the gastric cavity, based on the use of mucoadhesive films. Mucoadhesive properties of four formulations, with different chemical components and concentration, are evaluated by using both in vitro and ex vivo test benches on porcine stomach samples. Hydration of mucoadhesive films by contact with the aqueous mucous layer is analyzed by means of in vitro swelling tests, whereas optimal preloading conditions and adhesion performances, in terms of detachment force, supported weight and size are investigated ex vivo. Mucoadhesion is observed with all the four formulations. For a contact area of 113 mm2, the maximum normal and shear detachment forces withstood by the adhesive film are 2,6 N and 1 N respectively. These values grow up to 12,14 N and 4,5 N when the contact area increases to 706 mm2. Lifetime of the bonding on the inner side of the stomach wall was around two hours. Mucoadhesive anchoring represents a fully biocompatible and safe approach to deploy multiple assistive surgical tools on mucosal tissues by minimizing the number of access ports. This technique has been quantitatively assessed ex vivo for anchoring on the inner wall of the gastric cavity or in gastroscopic surgery. By properly varying the chemical formulation, this approach can be extended to other cavities of the human body.