Medical Imaging of Microrobots: Toward In Vivo Applications

Medical microrobots (MRs) have been demonstrated for a variety of non-invasive biomedical applications, such as tissue engineering, drug delivery, and assisted fertilization, among others. However, most of these demonstrations have been carried out in in vitro settings and under optical microscopy, being significantly different from the clinical practice. Thus, medical imaging techniques are required for localizing and tracking such tiny therapeutic machines when used in medical-relevant applications. This review aims at analyzing the state of the art of microrobots imaging by critically discussing the potentialities and limitations of the techniques employed in this field. Moreover, the physics and the working principle behind each analyzed imaging strategy, the spatiotemporal resolution, and the penetration depth are thoroughly discussed. The paper deals with the suitability of each imaging technique for tracking single or swarms of MRs and discusses the scenarios where contrast or imaging agent's inclusion is required, either to absorb, emit, or reflect a determined physical signal detected by an external system. Finally, the review highlights the existing challenges and perspective solutions which could be promising for future in vivo applications

Enzyme Powered Nanomotors Towards Biomedical Applications

[eng] The advancements in nanotechnology enabled the development of new diagnostic tools and drug delivery systems based on nanosystems, which offer unique features such as large surface area to volume ratio, cargo loading capabilities, increased circulation times, as well as versatility and multifunctionality. Despite this, the majority of nanomedicines do not translate into clinics, in part due to the biological barriers present in the body. Synthetic nano- and micromotors could be an alternative tool in nanomedicine, as the continuous propulsion force and potential to modulate the medium may aid tissue penetration and drug diffusion across biological barriers. Enzyme-powered motors are especially interesting for biomedical applications, owing to their biocompatibility and use of bioavailable substrates as fuel for propulsion. This thesis aims at exploring the potential applications of urease-powered nanomotors in nanomedicine. In the first work, we evaluated these motors as drug delivery systems. We found that active urease- powered nanomotors showed active motion in phosphate buffer solutions, and enhanced in vitro drug release profiles in comparison to passive nanoparticles. In addition, we observed that the motors were more efficient in delivering drug to cancer cells and caused higher toxicity levels, due to the combination of boosted drug release and local increase of pH produced by urea breakdown into ammonia and carbon dioxide. One of the major goals in nanomedicine is to achieve localized drug action, thus reducing side-effects. A commonly strategy to attain this is the use moieties to target specific diseases. In our second work, we assessed the ability of urease-powered nanomotors to improve the targeting and penetration of spheroids, using an antibody with therapeutic potential. We showed that the combination of active propulsion with targeting led to a significant increase in spheroid penetration, and that this effect caused a decrease in cell proliferation due to the antibody’s therapeutic action. Considering that high concentrations of nanomedicines are required to achieve therapeutic efficiency; in the third work we investigated the collective behavior of urease-powered nanomotors. Apart from optical microscopy, we evaluated the tracked the swarming behavior of the nanomotors using positron emission tomography, which is a technique widely used in clinics, due to its noninvasiveness and ability to provide quantitative information. We showed that the nanomotors were able to overcome hurdles while swimming in confined geometries. We observed that the nanomotors swarming behavior led to enhanced fluid convection and mixing both in vitro, and in vivo within mice’s bladders. Aiming at conferring protecting abilities to the enzyme-powered nanomotors, in the fourth work, we investigated the use of liposomes as chassis for nanomotors, encapsulating urease within their inner compartment. We demonstrated that the lipidic bilayer provides the enzymatic engines with protection from harsh acidic environments, and that the motility of liposome-based motors can be activated with bile salts. Altogether, these results demonstrate the potential of enzyme-powered nanomotors as nanomedicine tools, with versatile chassis, as well as capability to enhance drug delivery and tumor penetration. Moreover, their collective dynamics in vivo, tracked using medical imaging techniques, represent a step-forward in the journey towards clinical translation.[spa] Recientes avances en nanotecnología han permitido el desarrollo de nuevas herramientas para el diagnóstico de enfermedades y el transporte dirigido de fármacos, ofreciendo propiedades únicas como encapsulación de fármacos, el control sobre la biodistribución de estos, versatilidad y multifuncionalidad. A pesar de estos avances, la mayoría de nanomedicinas no consiguen llegar a aplicaciones médicas reales, lo cual es en parte debido a la presencia de barreras biológicas en el organismo que limitan su transporte hacia los tejidos de interés. En este sentido, el desarrollo de nuevos micro- y nanomotores sintéticos, capaces de autopropulsarse y causar cambios locales en el ambiente, podrían ofrecer una alternativa para la nanomedicina, promoviendo una mayor penetración en tejidos de interés y un mejor transporte de fármacos a través de las barreras biológicas. En concreto, los nanomotores enzimáticos poseen un alto potencial para aplicaciones biomédicas gracias a su biocompatibilidad y a la posibilidad de usar sustancias presentes en el organismo como combustible. Los trabajos presentados en esta tesis exploran el potenical de nanomotores, autopropulsados mediante la enzima ureasa, para aplicaciones biomédicas, y investigan su uso como vehículos para transporte de fármacos, su capacidad para mejorar penetración de tejidos diana, su versatilidad y movimiento colectivo. En conjunto, los resultados presentados en esta tesis doctoral demuestran el potencial del uso de nanomotores autopropulsados mediante enzimas como herramientas biomédicas, ofreciendo versatilidad en su diseño y una alta capacidad para promover el transporte de fármacos y la penetración en tumores. Por último, su movimiento colectivo observado in vivo mediante técnicas de imagen médicas representan un significativo avance en el viaje hacia su aplicación en medicina

Magnetically Driven Micro and Nanorobots

Manipulation and navigation of micro and nanoswimmers in different fluid environments can be achieved by chemicals, external fields, or even motile cells. Many researchers have selected magnetic fields as the active external actuation source based on the advantageous features of this actuation strategy such as remote and spatiotemporal control, fuel-free, high degree of reconfigurability, programmability, recyclability, and versatility. This review introduces fundamental concepts and advantages of magnetic micro/nanorobots (termed here as "MagRobots") as well as basic knowledge of magnetic fields and magnetic materials, setups for magnetic manipulation, magnetic field configurations, and symmetry-breaking strategies for effective movement. These concepts are discussed to describe the interactions between micro/nanorobots and magnetic fields. Actuation mechanisms of flagella-inspired MagRobots (i.e., corkscrew-like motion and traveling-wave locomotion/ciliary stroke motion) and surface walkers (i.e., surface-assisted motion), applications of magnetic fields in other propulsion approaches, and magnetic stimulation of micro/nanorobots beyond motion are provided followed by fabrication techniques for (quasi)spherical, helical, flexible, wire-like, and biohybrid MagRobots. Applications of MagRobots in targeted drug/gene delivery, cell manipulation, minimally invasive surgery, biopsy, biofilm disruption/eradication, imaging-guided delivery/therapy/surgery, pollution removal for environmental remediation, and (bio)sensing are also reviewed. Finally, current challenges and future perspectives for the development of magnetically powered miniaturized motors are discussed

Improved kinematic models for two-link helical micro/nano-swimmers

Accurate prediction of the three-dimensional trajectories of micro/nano-swimmers is a key element as to achieve high precision motion control in therapeutic applications. Rigid-body kinematics of such robotic systems is dominated by viscous forces. The induced flow field around a two-link swimmer is investigated with a validated computational fluid dynamics (CFD) model. Force-free-swimming constraints are employed in order to simulate motion of bacteria-like swimmers in viscous medium. The fluid resistance exerted on the body of the swimmer is quantified by an improved resistance matrix, which is embedded in a validated resistive force theory (RFT) model, based on complex-impedance approach. Parametric studies confirmed that the hydrodynamic interaction between body and tail are of great importance in predicting the trajectories for such systems

Analysis and Modeling of Magnetized Microswimmers: Effects of Geometry and Magnetic Properties

In recent years, much effort has been placed on development of microscale devices capable of propulsion in fluidic environments. These devices have numerous possible applications in biomedicine, microfabrication and sensing. One type of these devices that has drawn much attention among researchers is magnetic microswimmers--artificial microrobots that propel in fluid environments by being actuated using rotating external magnetic fields. This dissertation highlights our contribution to this class of microrobots. We address issues regarding fabrication difficulties arising from geometric complexities as well as issues pertaining to the controllability and adaptability of microswimmers.The majority of research in this field focuses on utilization of flexible or achiral geometries as inspired by microbiological organisms such as sperm and bacteria. Here, we set forth the minimum geometric requirements for feasible designs and demonstrate that neither flexibility nor chirality is required, contrary to biomimetic expectations. The physical models proposed in this work are generally applicable to any geometry and are capable of predicting the swimming behavior of artificial microswimmers with permanent dipoles. Through these models, we explain the wobbling phenomena, reported by experimentalists. Our model predicts the existence of multiple stable solutions under certain conditions. This leads to the realization that control strategies can be improved by adjusting the angle between the applied magnetic field and its axis of rotation. Furthermore, we apply our model to helical geometries which encompass the majority of magnetic microswimmers. We demonstrate the criterion for linear velocity-frequency response and minimization of wobbling motion. One approach to improve the adaptability of swimmers to various environments is to use modular units that can dynamically assemble and disassemble on-site. We propose a model to explain the docking process which informs strategies for successful assemblies. Most studies conducted so far are to elucidate permanent magnetic swimmers, but the literature is lacking on analysis of swimmers made of soft ferromagnetic materials. In this work, we develop a model for soft-magnetic microswimmers in the saturation regime in order to predict the swimming characteristics of these types of swimmers and compare to those of hard-magnetic swimmers

Innovative designs and applications of Janus micromotors with (photo)-catalytic and magnetic motion

El objetivo principal de esta Tesis Doctoral es el diseño y desarrollo de micromotores Janus biocompatibles y su aplicación en ámbitos relevantes de la salud y de la protección medioambiental. Los micromotores Janus son dispositivos en la microescala autopropulsados que tienen al menos dos regiones en su superficie con diferentes propiedades físicas y químicas, lo que les convierte en una clase distintiva de materiales que pueden combinar características ópticas, magnéticas y eléctricas en una sola entidad. Como la naturaleza del micromotor Janus -el dios romano de las dos caras- los objetivos de esta Tesis Doctoral presentan naturaleza dual y comprenden desarrollos de química fundamental y de química aplicada. En efecto, por una parte, el objetivo central aborda el diseño, síntesis y ensamblaje, así como la caracterización de micromotores Janus poliméricos propulsados por mecanismos (foto)-catalíticos y/o accionados por campos magnéticos. Por otra parte, el objetivo central implica la aplicación de los micromotores desarrollados para resolver desafíos sociales relevantes en los ámbitos químico-analítico, biomédico y ambiental. Partiendo de estas premisas, en la primera parte de la Tesis Doctoral, se sintetizaron micromotores Janus de policaprolactona propulsados químicamente integrando nanomateriales para el diseño de sensores móviles para la detección selectiva de endotoxinas bacterianas. De esta forma, el movimiento autónomo del micromotor mejora la mezcla de fluidos y la eficacia de las reacciones implicadas permitiendo detectar el analito en pocos minutos, incluso en muestras viscosas y medios donde la agitación no es posible. Además, esta autopropulsión es altamente compatible con su empleo en formatos ultra-miniaturizados para el desarrollo de futuros dispositivos portátiles en el marco de la tecnología point of care para aplicaciones clínicas y agroalimentarias. Con el fin de incrementar su biocompatibilidad para aplicaciones in vivo, en una segunda etapa de la Tesis Doctoral, se diseñaron micromotores Janus con propulsión autónoma utilizando luz visible para la eliminación de toxinas relevantes en procesos inflamatorios. El fenómeno autopropulsivo del micromotor y su capacidad de interacción con agentes tóxicos condujo a metodologías más rápidas y eficaces infiriéndose un futuro prometedor de estos micromotores para el tratamiento del shock séptico o intoxicación. En una tercera etapa, se sintetizaron micromotores propulsados por campos magnéticos. Estos micromotores utilizan una aproximación elegante de propulsión, exenta del empleo de combustibles químicos tóxicos como sucede en la propulsión catalítica y, en consecuencia, biocompatible. Asimismo, este mecanismo propulsivo permite controlar e incluso programar su trayectoria para aplicaciones que requieran de un guiado y de un control preciso de esta. De manera específica, estos micromotores han sido aplicados en esta Tesis Doctoral para la liberación controlada de fármacos en el tratamiento de cáncer pancreático y como elementos de remediación ambiental en la eliminación de agentes nerviosos en aguas contaminadas

Microdevices and Microsystems for Cell Manipulation

Microfabricated devices and systems capable of micromanipulation are well-suited for the manipulation of cells. These technologies are capable of a variety of functions, including cell trapping, cell sorting, cell culturing, and cell surgery, often at single-cell or sub-cellular resolution. These functionalities are achieved through a variety of mechanisms, including mechanical, electrical, magnetic, optical, and thermal forces. The operations that these microdevices and microsystems enable are relevant to many areas of biomedical research, including tissue engineering, cellular therapeutics, drug discovery, and diagnostics. This Special Issue will highlight recent advances in the field of cellular manipulation. Technologies capable of parallel single-cell manipulation are of special interest

Modeling and simulation of uni- and multi-flagellar bacterial locomotion in a viscous fluid

Flagellated bacteria categorized as microorganisms, play vital roles in human life such that their beneficial and detrimental effects on different aspects of the ecosystem are indisputable. Flagellated bacteria propel themselves in fluids by rotating one or more flexible filaments, known as flagella, driven by independent flagellar motors. Depending on the rotation direction of the motors and handedness of the helical filaments, the flagella either pull or push the cell body. Entrapment of swimming bacteria near surfaces, observed in some species, may lead to biological processes such as biofilm formation and wound infection. Previous experimental and numerical studies of bacterial locomotion have illustrated that several behaviors exhibited by the bacteria have roots in hydrodynamic interactions between the bacteria components and the surrounding fluid. In this thesis, we numerically study flagellated bacterial locomotion in bounded and unbounded spaces. The physical properties of the model bacteria in this study are described based on experimental data available for various species of uni-, bi-, and multiflagellated bacteria. Specifically, we choose Vibrio alginolyticus, Magnetococcus marinus and Escherichia coli to focus on their motility to shed light on some of the unique behavior observed in each one. Depending on the species, the model bacteria have either a spherical or a spherocylindrical cell body and the flexible flagellar filaments are connected to the cell body membrane directly or via very flexible straight hooks. The flagella are independently driven by either constant or variable torque motors. Despite a similar flagellar structure, uni- and multiflagellated bacteria employ different mechanisms to swim on straight trajectories or reorient. Here, we use the boundary element method (BEM) and the Kirchhoff rod model to develop a comprehensive elastohydrodynamic framework in order to model the motility of uni- and multiflagellated bacteria in a Newtonian viscous fluid. For this purpose, the boundary integral equations (BIE) are numerically evaluated over the cell body surface and along the flagella which are described by distributions of regularized Stokeslets and Rotlets. By assuming that the flagella are inextensible and unshearable, the linear theory of elasticity is used to estimate the internal moments along the flagella. Adding the hydrodynamics and elasticity equations to the total force/torque balance and kinematic equations leads to a system of linear equations which are solved to find the velocities and update the swimmer configuration accordingly. Motivated by experimental observations of Vibrio alginolyticus locomotion in which it is shown that there is an interesting correlation between the near-surface entrapment of bacteria and the concentration of certain ions in the swimming medium, we numerically investigate its motility in different concentrations of NaCl. Our simulations demonstrate that changing the concentration of NaCl in the swimming fluid affects the tendency of pusher-mode bacteria to remain near the surfaces by altering the averaged swimming speed and inducing the different degrees of deformations along the flagellum. In addition to the ion concentration, our results indicate the flagellum/hook stiffness, the flagellar motor torque, and the cell body aspect ratio may affect whether the uniflagellated model bacterium escapes from the surface or becomes trapped in circular orbits. By simulating the locomotion of a bi-flagellated model bacterium with a spherical cell body, one puller, and one pusher flagellum, we show that the bacteria with such configuration mainly swim along double helical trajectories. Comparing the properties of the obtained trajectories with the Magnetococcus marinus's trajectories measured experimentally, indicate that this species has likely puller-pusher configuration. Varying the stiffness, orientations, or positions of the flagella significantly changes the swimming characteristics. Notably, when either the applied torque to the pusher flagellum is higher than a critical value and/or its stiffness is lower than a critical stiffness, the pusher flagellum exhibits overwhirling motion, resulting in a more complicated swimming style and a lower swimming speed. For a moderate flagellum stiffness, the swimming speed is insensitive to the rest orientation of the flagella over a wide range of orientation angles because the flagella deform to maintain alignment with the swimming direction. Numerical investigation of multiflagellated bacteria locomotion in unbounded fluid indicates that the arrangement of the flagella on the cell body provides no advantages in the average swimming speeds of bacteria. However, the trajectory of the bacteria could be either relatively straight or double helical trajectory depending on the degree of asymmetry that exists in the distribution of the flagella. Our results indicate that the multiflagellated bacteria in the ``run'' state may have several stable swimming modes in which the swimming properties such as speeds and trajectories could be different. The tumbling event, stopping of the flagellar motor, and interaction with other bacteria are likely some reasons which cause the bacteria to switch between the different modes. High viscous torque due to the presence of a no-slip boundary slightly changes the swimming properties of the multiflagellated bacteria such as bundling time, the translational and angular speeds. Remarkably, the flagella arrangement is one of the key factors determining how the swimming properties vary in response to the presence of a surface