Direct and rapid intracellular delivery of a functional Cas9/sgRNA complex using ultrasound-powered nanomotors is reported. The Cas9/sgRNA complex is loaded onto the nanomotor surface through a reversible disulfide linkage. A 5 min ultrasound treatment enables the Cas9/sgRNA-loaded nanomotors to directly penetrate through the plasma membrane of GFP-expressing B16F10 cells. The Cas9/sgRNA is released inside the cells to achieve highly effective GFP gene knockout. The acoustic Cas9/sgRNA-loaded nanomotors display more than 80 % GFP knockout within 2 h of cell incubation compared to 30 % knockout using static nanowires. More impressively, the nanomotors enable highly efficient knockout with just 0.6 nm of the Cas9/sgRNA complex. This nanomotor-based intracellular delivery method thus offers an attractive route to overcome physiological barriers for intracellular delivery of functional proteins and RNAs, thus indicating considerable promise for highly efficient therapeutic applications.

Vaccination represents one of the most effective means of preventing infectious disease. In order to maximize the utility of vaccines, highly potent formulations that are easy to administer and promote high patient compliance are desired. In the present work, a biomimetic self-propelling micromotor formulation is developed for use as an oral antivirulence vaccine. The propulsion is provided by a magnesium-based core, and a biomimetic cell membrane coating is used to detain and neutralize a toxic antigenic payload. The resulting motor toxoids leverage their propulsion properties in order to more effectively elicit mucosal immune responses. After demonstrating the successful fabrication of the motor toxoids, their uptake properties are shown in vitro. When delivered to mice via an oral route, it is then confirmed that the propulsion greatly improves retention and uptake of the antigenic material in the small intestine in vivo. Ultimately, this translates into markedly elevated generation of antibody titers against a model toxin. This work provides a proof-of-concept highlighting the benefits of active oral delivery for vaccine development, opening the door for a new set of applications in which biomimetic motor technology can provide significant benefits.

Tremendous progress has been made during the past decade toward the design of nano/micromotors with high biocompatibility, multifunctionality, and efficient propulsion in biological fluids, which collectively have led to the initial investigation of in vivo biomedical applications of these synthetic motors. Despite these recent advances in micromotor designs and mechanistic research, significant effort is needed to develop appropriate formulations of micromotors to facilitate their in vivo administration and thus to better test their in vivo applicability. Herein, we present a micromotor pill and demonstrate its attractive use as a platform for in vivo oral delivery of active micromotors. The micromotor pill is comprised of active Mg-based micromotors dispersed uniformly in the pill matrix, containing inactive (lactose/maltose) excipients and other disintegration-aiding (cellulose/starch) additives. Our in vivo studies using a mouse model show that the micromotor pill platform effectively protects and carries the active micromotors to the stomach, enabling their release in a concentrated manner. The micromotor encapsulation and the inactive excipient materials have no effects on the motion of the released micromotors. The released cargo-loaded micromotors propel in gastric fluid, retaining the high-performance characteristics of in vitro micromotors while providing higher cargo retention onto the stomach lining compared to orally administrated free micromotors and passive microparticles. Furthermore, the micromotor pills and the loaded micromotors retain the same characteristics and propulsion behavior after extended storage in harsh conditions. These results illustrate that combining the advantages of traditional pills with the efficient movement of micromotors offer an appealing route for administrating micromotors for potential in vivo biomedical applications.

tract delivery, [10] in vivo drug or antigen delivery, [11,12] and collective and dynamic gastric delivery via micromotor pills. [13] While Mg micromotors have been widely tested for dynamic in vivo delivery applications, their ability to manipulate, carry, and transport living cells has not been explored.Along the line of micromotor design, significant progress has been made recently toward creating biohybrid micromotors that combine cellular components and synthetic micro/nanoscale materials. Such cell-based micromotors offer considerable promise for diverse in vivo biomedical applications owing to their biocompatibility and biological functionality of the cellular component. [14] Live cells can thus be integrated with artificial substrates to produce functional biohybrid devices that possess new and improved capabilities. Such cell-based micromotors can be classified in two types. The first one relies on the intrinsic motility of live cells, such as spermatozoa, [15] bacteria, [16] and cardiomyocytes, [17] for transporting artificial material payloads. The microorganisms thus act as engines to form active biohybrid swimming systems powered by cellular actuation. The second type consists of cell-based materials, such as cell membranes, and synthetic micromotors that provide the motion. Such combination of micromotors with cell components confers the micromotor with cell-like properties [18,19] and improved biofunctionality. [20] Besides cell membrane-coated micromotors, various types of cells have also been used in biomedical applications due to their large drug loading capacity, [21] natural homing tendency toward inflammation sites, [22] and easy genetic engineering for gene delivery. [23] The combination of intact cells and engineered motors has resulted in several cellbased biohybrid micromotor designs for diverse applications, including stem cell-based motors for drug delivery, [24] red blood cell-motors for on-demand cargo delivery, [25] and NIH 3T3 cellbased motors for precise control and patterning. [26] Although such cell-based motors have demonstrated clear advantages as drug delivery carriers, it will be particularly interesting to explore the possibility of integrating intact live cells with biocompatible and biodegradable artificial micromotors, such as the Mg-based micromotors, to form a biohybrid motor system, which can potentially be applied for in vivo operations. Magnesium (Mg)-based micromotors are combined with live macrophage (MΦ) cells to create a unique MΦ-Mg biohybrid motor system. The resulting biomotors possess rapid propulsion ability stemming from the Mg micromotors and the biological functions provided by the live MΦ cell. To prepare the biohybrid motors, Mg microparticles coated with titanium dioxide and poly(l-lysine) (PLL) layers are incubated with live MΦs at low temperature. The formation of such biohybrid motors depends on the relative size of the MΦs and Mg particles, with the MΦ swallowing up Mg particles smaller than 5 µm. The experimental results and numerical simulations ...

As the most common nutritional disorder, iron deficiency represents a major public health problem with broad impacts on physical and mental development. However, treatment is often compromised by low iron bioavailability and undesired side effects. Here, we report on the development of active mineral delivery vehicles using Mg-based micromotors, which can autonomously propel in gastrointestinal fluids, aiding in the dynamic delivery of minerals. Iron and selenium are combined as a model mineral payload in the micromotor platform. We demonstrate the ability of our mineral-loaded micromotors to replenish iron and selenium stores in an anemic mouse model after 30 days of treatment, normalizing hematological parameters such as red blood count, hemoglobin, and hematocrit. Additionally, the micromotor platform exhibits no toxicity after the treatment regimen. This proof-of-concept study indicates that micromotor-based active delivery of mineral supplements represents an attractive approach toward alleviating nutritional deficiencies.