Biohybrid Systems Research Articles

Propulsion and Chemotaxis in Bacteria-Driven Microswimmers.

Despite the large body of experimental work recently on biohybrid microsystems, few studies have focused on theoretical modeling of such systems, which is essential to understand their underlying functioning mechanisms and hence design them optimally for a given application task. Therefore, this study focuses on developing a mathematical model to describe the 3D motion and chemotaxis of a type of widely studied biohybrid microswimmer, where spherical microbeads are driven by multiple attached bacteria. The model is developed based on the biophysical observations of the experimental system and is validated by comparing the model simulation with experimental 3D swimming trajectories and other motility characteristics, including mean squared displacement, speed, diffusivity, and turn angle. The chemotaxis modeling results of the microswimmers also agree well with the experiments, where a collective chemotactic behavior among multiple bacteria is observed. The simulation result implies that such collective chemotaxis behavior is due to a synchronized signaling pathway across the bacteria attached to the same microswimmer. Furthermore, the dependencies of the motility and chemotaxis of the microswimmers on certain system parameters, such as the chemoattractant concentration gradient, swimmer body size, and number of attached bacteria, toward an optimized design of such biohybrid system are studied. The optimized microswimmers would be used in targeted cargo, e.g., drug, imaging agent, gene, and RNA, transport and delivery inside the stagnant or low‐velocity fluids of the human body as one of their potential biomedical applications.

Combined influence of biophysical and biochemical cues on maintenance and proliferation of hematopoietic stem cells.

Homeostasis of hematopoietic stem and progenitor cells (HSPC) is controlled by a combination of biochemical and biophysical environmental cues in the bone marrow (BM) niche, where a tight balance of quiescence and proliferation of HSPC is maintained. Specifically, alongside soluble factors and extracellular matrix (ECM) proteins, spatial confinement and ECM stiffness have been recognized to be critical for regulation of HSPC fate. Here we employ a modular, glycosaminoglycan (GAG)-based biohybrid hydrogel system to balance proliferation of human HSPC and maintenance of quiescent hematopoietic stem cells (HSC) through simultaneous regulation of exogenous biochemical and biophysical cues. Our results demonstrate that HSPC respond to increased spatial confinement with lowered proliferation and cell cycling, which results in higher frequency of quiescent LTC-IC (long-term culture initiating cells), while GAG-rich 3D environments further support maintenance of the cells.

Bioelectronic Devices: Cell Generator: A Self‐Sustaining Biohybrid System Based on Energy Harvesting from Engineered Cardiac Microtissues (Adv. Funct. Mater. 20/2017)

Bioelectronic Devices: Cell Generator: A Self‐Sustaining Biohybrid System Based on Energy Harvesting from Engineered Cardiac Microtissues (Adv. Funct. Mater. 20/2017)

Inorganic/whole-cell biohybrid photocatalyst for highly efficient hydrogen production from water

To obtain a clean hydrogen production system, we have developed an inorganic-bio hybrid photocatalyst system based on the combination of anatase TiO2, methylviologen (MV) as an electron mediator, and a whole-cell biocatalyst consisting of [FeFe]-hydrogenase and maturase gene-harboring recombinant Escherichia coli; however, the apparent quantum yield at 300nm (AQY300) for hydrogen production was low (0.3%). The system consists of a two-step reaction: (1) photocatalytic MV reduction by TiO2, and (2) hydrogen production with reduced MV using a biocatalyst. The enhancement of step 1 under biocatalyst-friendly conditions was investigated in an attempt to further improve the reaction efficiency. Among the condition tested, the use of 100mM Tris-HCl (pH 7), 150mM NaCl, and 5% (v/v) glycerol with P-25 TiO2 especially enhanced the step 1 reaction by a 300-fold increase in the MV reduction rate compared with previously tested reaction condition (100mM Tris-HCl (pH 7), 150mM NaCl, 5% (v/v) glycerol, and 100mM ascorbate with anatase TiO2). Under the enhanced step 1 reaction, AQY300 and AQY350 for photocatalytic MV reduction reached 60.8% and 52.2%, respectively. The enhanced step 1 reaction thus significantly improved the overall photocatalytic hydrogen productivity of the hybrid system and AQY300 and AQY350 reached 26.4% and 31.2%, respectively. The inorganic-whole-cell biohybrid system can therefore provide noble metal-free, efficient, and clean hydrogen production.

Cell Generator: A Self‐Sustaining Biohybrid System Based on Energy Harvesting from Engineered Cardiac Microtissues

Biohybrid soft robotic devices present unique advantages for designing biologically active machines that can dynamically sense and interact with complex bioelectrical signals. Here, a controllable cell‐based machine is developed that harvests energy from arrays of beating cardiomyocytes to generate electricity for biomedical microscale robotic applications. The “Cell Generator” device is based on an array of piezoelectric microcantilevers wrapped with 3D patterned cardiac cells. Spontaneous contraction of the engineered cardiac constructs provides the source of mechanical energy for electricity generation. It is demonstrated that a single “Cell Generator” unit with 40 cantilevers can output peak voltages of ≈70 mV, and a larger array of 540 cantilevers can directly generate a pulsed output as high as ≈1 V. When integrated with an electrical rectification and storage circuit, it is further shown that the “Cell Generator” can provide functional outputs and work as a self‐powered neural stimulator to evoke action potentials in cultured neuronal networks. This demonstration of “Cell Generator” technology provides an innovative perspective of exploiting live biological powering system on biomedical microscale robotic devices in the human body.

Cyborgian Material Design for Solar Fuel Production: The Emerging Photosynthetic Biohybrid Systems.

Photosynthetic biohybrid systems (PBSs) combine the strengths of inorganic materials and biological catalysts by exploiting semiconductor broadband light absorption to capture solar energy and subsequently transform it into valuable CO2-derived chemicals by taking advantage of the metabolic pathways in living organisms. In this work, we first traverse through a brief history of recent PBSs, demonstrating the modularity and diversity of possible architectures to rival and, in many cases, surpass the performance of chemistry or biology alone before envisioning the future of these hybrid systems, opportunities for improvement, and its role in sustainable living here on earth and beyond.

Multicellular Biohybrid Materials: Probing the Interplay of Cells of Different Types Precisely Positioned and Constrained on 3D Wireframe-Like Microstructures.

Driven by the unbroken miniaturization trend in microtechnology, the development of smaller, yet reliable and efficient, highly integrated microsystems can benefit from inherent capabilities of biological cells. In particular, by featuring multiple types of cells, biohybrid systems exhibiting self-contained sensing and actuation capabilities can be conceived. To ensure the proper functioning of such multicellular biohybrid systems, the intended cell arrangement needs to be maintained over time. Microscaffolds designed for this purpose should therefore selectively guide or hinder cell migration. However, the basic cell-structure interactions governing the cell migration and extension processes are not yet fully understood. This paper explores these interactions and proposes a method for the fabrication of advanced multicellular biohybrid materials. The method is based on wireframe-like 3D microstructures onto which several types of cells are successfully positioned and arranged by optical manipulation. Experiments exploring cell dynamics reveal geometry-dependent maximal migration and extension distances. Microscaffolds designed on the basis of these characteristics can guide cell migration, trigger structure-contained cell growth, and maintain a predetermined cell arrangement. The methods reported herein therefore provide insight into cell assembly and migration on 3D microscaffolds, which is an essential early step towards advanced multicellular biohybrid materials.

Biohybrid Membrane Systems and Bioreactors as Tools for In Vitro Drug Testing.

In drug development, in vitro human model systems are absolutely essential prior to the clinical trials, considering the increasing number of chemical compounds in need of testing, and, keeping in mind that animals cannot predict all the adverse human health effects and reactions, due to the species-specific differences in metabolic pathways. The liver plays a central role in the clearance and biotransformation of chemicals and xenobiotics. In vitro liver model systems by using highly differentiated human cells could have a great impact in preclinical trials. Membrane biohybrid systems constituted of human hepatocytes and micro- and nano-structured membranes, represent valuable tools for studying drug metabolism and toxicity. Membranes act as an extracellular matrix for the adhesion of hepatocytes, and compartmentalise them in a well-defined physical and chemical microenvironment with high selectivity. Advanced 3-D tissue cultures are furthermore achieved by using membrane bioreactors (MBR), which ensure the continuous perfusion of cells protecting them from shear stress. MBRs with different configurations allow the culturing of cells at high density and under closely monitored high perfusion, similarly to the natural liver. These devices that promote the long-term maintenance and differentiation of primary human hepatocytes with preserved liver specific functions can be employed in drug testing for prolonged exposure to chemical compounds and for assessing repeated-dose toxicity. The use of primary human hepatocytes in MBRs is the only system providing a faster and more cost-effective method of analysis for the prediction of in vitro human drug metabolism and enzyme induction alternative and/or complementary to the animal experimentation. In this paper, in vitro models for studying drug metabolism and toxicity as advanced biohybrid membrane systems and MBRs will be reviewed.

Advances in Hybrid Polymer-Based Materials for Sustained Drug Release

The use of biomaterials composed of organic pristine components has been successfully described in several purposes, such as tissue engineering and drug delivery. Drug delivery systems (DDS) have shown several advantages over traditional drug therapy, such as greater therapeutic efficacy, prolonged delivery profile, and reduced drug toxicity, as evidenced byin vitroandin vivostudies as well as clinical trials. Despite that, there is no perfect delivery carrier, and issues such as undesirable viscosity and physicochemical stability or inability to efficiently encapsulate hydrophilic/hydrophobic molecules still persist, limiting DDS applications. To overcome that, biohybrid systems, originating from the synergistic assembly of polymers and other organic materials such as proteins and lipids, have recently been described, yielding molecularly planned biohybrid systems that are able to optimize structures to easily interact with the targets. This work revised the biohybrid DDS clarifying their advantages, limitations, and future perspectives in an attempt to contribute to further research of innovative and safe biohybrid polymer-based system as biomaterials for the sustained release of active molecules.

Connecting quantum dots with enzymes: mediator-based approaches for the light-directed read-out of glucose and fructose oxidation.

The combination of the biocatalytic features of enzymes with the unique physical properties of nanoparticles in a biohybrid system provides a promising approach for the development of advanced bioelectrocatalytic devices. This study describes the construction of photoelectrochemical signal chains based on CdSe/ZnS quantum dot (QD) modified gold electrodes as light switchable elements, and low molecular weight redox molecules for the combination with different biocatalysts. Photoelectrochemical and photoluminescence experiments verify that electron transfer can be achieved between the redox molecules hexacyanoferrate and ferrocene, and the QDs under illumination. Since for both redox mediators a concentration dependent photocurrent change has been found, light switchable enzymatic signal chains are built up with fructose dehydrogenase (FDH) and pyrroloquinoline quinone-dependent glucose dehydrogenase ((PQQ)GDH) for the detection of sugars. After immobilization of the enzymes at the QD electrode the biocatalytic oxidation of the substrates can be followed by conversion of the redox mediator in solution and subsequent detection at the QD electrode. Furthermore, (PQQ)GDH has been assembled together with ferrocenecarboxylic acid on top of the QD electrode for the construction of a funtional biohybrid architecture, showing that electron transfer can be realized from the enzyme over the redox mediator to the QDs and subsequently to the electrode in a completely immobilized fashion. The results obtained here do not only provide the basis for light-switchable biosensing and bioelectrocatalytic applications, but may also open the way for self-driven point-of-care systems by combination with solar cell approaches (power generation at the QD electrode by enzymatic substrate consumption).

Cell-Based Biohybrid Drug Delivery Systems: The Best of the Synthetic and Natural Worlds.

The goal of drug delivery is to deliver therapeutics to the site of disease while reducing unwanted side effects. In recent years, a diverse variety of synthetic nano and microparticles have been developed as drug delivery systems. The success of these systems for drug delivery lies in their ability to overcome biological barriers such as the blood-brain barrier, to evade immune clearance and avoid nonspecific biodistribution. This Review provides an overview of recent advances in the design of biohybrid drug delivery systems, which combine cells with synthetic systems to overcome some of these biological hurdles. Examples include eukaryotic cells, such as stem cells, red blood cells, immune cells, platelets, and cancer cells that are used to carry drug-loaded synthetic particles. Synthetic particles can also be cloaked with naturally derived cell membranes and thereby evade immune clearance, exhibit prolonged systemic circulation, and target specific tissues by capitalizing on the interaction/homing tendency of certain cells and their membrane components to particular tissues. Different designs of cell-based biohybrid systems and their applications, as well as their promise and limitations, are discussed herein.

Investigation of Collective Behaviour and Electrocommunication in the Weakly Electric Fish, Mormyrus rume, through a biomimetic Robotic Dummy Fish

A robotic fish has been developed to create a mixed bio-hybrid system made up of weakly electric fish and a mobile dummy fish. Weakly electric fish are capable of interacting with each other via sequences of self-generated electric signals during electrocommunication. Here we present the design of an artificial dummy fish, which is subsequently tested in behavioural experiments. The robot consists of two parts: a flexible tail that can move at different frequencies and amplitudes, performing a carangiform oscillation, and a rigid head containing the motor for the tail oscillation. The dummy fish mimics the weakly electric fish Mormyrus rume in morphology, size and electric signal generation. In order to study electrical interactions, the dummy fish is equipped with ten electrodes that record electric signals of nearby real fish and generate electric dipole fields around itself that are similar to those produced by real fish in both waveform and sequence. Behavioural experiments demonstrate that the dummy fish is able to recruit both single individuals and groups of M. rume from a shelter into an exposed area. The development of an artificial dummy fish may help to understand fundamental aspects of collective behaviour in weakly electric fish and the properties necessary to initiate and sustain it in closed-loop feedback experiments based on electrocommunication.

A multidisciplinary approach to study the functional properties of neuron-like cell models constituting a living bio-hybrid system: SH-SY5Y cells adhering to PANI substrate

A living bio-hybrid system has been successfully implemented. It is constituted by neuroblastic cells, the SH-SY5Y human neuroblastoma cells, adhering to a poly-anyline (PANI) a semiconductor polymer with memristive properties. By a multidisciplinary approach, the biocompatibility of the substrate has been analyzed and the functionality of the adhering cells has been investigated. We found that the PANI films can support the cell adhesion. Moreover, the SH-SY5Y cells were successfully differentiated into neuron-like cells for in vitro applications demonstrating that PANI can also promote cell differentiation. In order to deeply characterize the modifications of the bio-functionality induced by the cell-substrate interaction, the functional properties of the cells have been characterized by electrophysiology and Raman spectroscopy. Our results confirm that the PANI films do not strongly affect the general properties of the cells, ensuring their viability without toxic effects on their physiology. Ascribed to the adhesion process, however, a slight increase of the markers of the cell suffering has been evidenced by Raman spectroscopy and accordingly the electrophysiology shows a reduction at positive stimulations in the cells excitability.

(Invited) Biohybrid Photoelectrodes Exploiting Effective Photosystem I - Electrode Contacts

Hybrid systems using the unique features of natural photosynthesis are increasingly in the focus of current research. Particularly the two protein complexes photosystem I and II (PS) of the oxygenic photosynthesis have attracted the attention of researchers to build up new solar energy-converting systems. In such biohybrid systems an efficient coupling of PS with the electrode is essential. Besides the light-to-current conversion, the PS may also be used for light-driven enzymatic reactions resulting in photobioelectrocatalysis. Different basic principles can be used by establishing communication between the PS and electrode surfaces. Here we want to avoid shuttle molecules and generate a surface allowing direct electron transfer (DET) between the excited protein and the electrode. This has been studied with photosystem 1 (PSI) from Thermosynechococcus elongates.For efficient photocurrent generation a arther oriented immobilisation would be desirable which is complicated by the rather high molecular weight and the complex surface properties of PSI. In this context we have been focusing on the directional assembly of PSI on highly conductive graphene electrodes. In order to adjust the surface properties different aromatic molecules have been applied onto the graphene [1]. The different Π-systems serve as an artificial scaffold harbouring functional groups which interplay with the PSI. Particularly important is the hydrophilic-hydrophobic balance on the surface. In conclusion, an effective and rather directed assembly of PSI and a large photocurrent density has been obtained. It has to be mentioned here that photocurrent generation occurs already at open circuit potential with now additional driving force applied. Depending on the modifier used an unidirectional photocurrent generation can be achieved by this approach. Via the artificially modified graphene surface we have been able to create biohybrid systems with an improved photocurrent output. Here not only adsorption can be used for the functional assembly but also covalent linkage of PSI to the surface. A systematic investigation on the protein assembly and the resulting photoelectrochemical properties will be presented. For this quartz crystal microbalance experiments, chopped light voltammetry, constant potential measurements, photoaction spectra and experiments with methyl viologen have been performed. Moreover it can be demonstrated that the performance of such PSI electrodes can be further improved. Here nanoparticles are additionally coupled to the graphene-PSI system in order to enhance current generation under illumination. [1] S. C. Feifel, K. R. Stieger, H. Lokstein, H. Lux, F. Lisdat, Journal of Materials Chemistry A 3 (23) (2015) 12188-12196 [2] S. C. Feifel, H. Lokstein, M. Hejazi, A. Zouni, F. Lisdat, Langmuir 31 (38) (2015) 10590-10598

Chemotaxis of bio-hybrid multiple bacteria-driven microswimmers

In this study, in a bio-hybrid microswimmer system driven by multiple Serratia marcescens bacteria, we quantify the chemotactic drift of a large number of microswimmers towards L-serine and elucidate the associated collective chemotaxis behavior by statistical analysis of over a thousand swimming trajectories of the microswimmers. The results show that the microswimmers have a strong heading preference for moving up the L-serine gradient, while their speed does not change considerably when moving up and down the gradient; therefore, the heading bias constitutes the major factor that produces the chemotactic drift. The heading direction of a microswimmer is found to be significantly more persistent when it moves up the L-serine gradient than when it travels down the gradient; this effect causes the apparent heading preference of the microswimmers and is the crucial reason that enables the seemingly cooperative chemotaxis of multiple bacteria on a microswimmer. In addition, we find that their chemotactic drift velocity increases superquadratically with their mean swimming speed, suggesting that chemotaxis of bio-hybrid microsystems can be enhanced by designing and building faster microswimmers. Such bio-hybrid microswimmers with chemotactic steering capability may find future applications in targeted drug delivery, bioengineering, and lab-on-a-chip devices.

Update on Renal Replacement Therapy: Implantable Artificial Devices and Bioengineered Organs.

Recent advances in the fields of artificial organs and regenerative medicine are now joining forces in the areas of organ transplantation and bioengineering to solve continued challenges for patients with end-stage renal disease. The waiting lists for those needing a transplant continue to exceed demand. Dialysis, while effective, brings different challenges, including quality of life and susceptibility to infection. Unfortunately, the majority of research outputs are far from delivering satisfactory solutions. Current efforts are focused on providing a self-standing device able to recapitulate kidney function. In this review, we focus on two remarkable innovations that may offer significant clinical impact in the field of renal replacement therapy: the implantable artificial renal assist device (RAD) and the transplantable bioengineered kidney. The artificial RAD strategy utilizes micromachining techniques to fabricate a biohybrid system able to mimic renal morphology and function. The current trend in kidney bioengineering exploits the structure of the native organ to produce a kidney that is ready to be transplanted. Although these two systems stem from different technological approaches, they are both designed to be implantable, long lasting, and free standing to allow patients with kidney failure to be autonomous. However, for both of them, there are relevant issues that must be addressed before translation into clinical use and these are discussed in this review.

Light‐Driven Highly Selective Conversion of CO2 to Formate by Electrosynthesized Enzyme/Cofactor Thin Film Electrode

The highly selective electrochemical reduction of carbon dioxide (CO2) is reported to formate (HCOO‐) at a compactly integrated bioelectrode. The enzymatic biocatalytic cathode is fabricated by single‐step electropolymerization of a multifunctional polydopamine film in which enzyme/cofactor couples are uniquely embedded. Interestingly, this thin biohybrid system of nanoscale thickness assures unprecedentedly prolonged catalytic enzyme stability for about two weeks. Mimicking the natural photosynthesis, combination with the photoanode utilizing the water oxidation, steadily produces formate at a faradaic efficiency of 99.18 ± 6.77% with little degradation at least for 1 d under 1 sun illumination and no external bias.

Experimental Approach Towards Long-Range Interactions from 1.6 to 13798 km Distances in Bio-Hybrid Systems

This work describes performed experiments on device-device and ope¬rator-device interactions at distances of >1 km, >100 km and >10000 km. Experimental setup involves several types of receiving sensors and transmitting optical generators as well as a group of human operators. We analyzed the structure of setup, establishing a connection between receiver and emitter, and multiple effects appeared. The experiments suggest a common character of operator- and device- interactions that point to possible ’neuro-quantum mechanisms’ underling both systems. This approach replicates and extends early experiments from 80x and 90x, and can be considered as a novel unconventional communication system.

Targeted Drug Delivery and Imaging Using Mobile Milli/Microrobots: A Promising Future Towards Theranostic Pharmaceutical Design.

Miniature untethered medical robots have been receiving growing attention due to technological advances in microactuation, microsensors, and microfabrication and have significant potential to reduce the invasiveness and improve the accessibility of medical devices into unprecedented small spaces inside the human body. In this review, we discuss therapeutic and diagnostic applications of untethered medical microrobots. Wirelessly controlled milli/microrobots with integrated sensors are revolutionizing micromanipulation based medical interventions and are enabling doctors to perform minimally invasive procedures not possible before. 3D fabrication technologies enabling milli/microrobot fabrication at the single cell scale are empowering high-resolution visual imaging and in vivo manipulation capabilities. Swallowable millirobots and injectabale ocular microrobots allow the gastric ulcer imaging, and performance of vitreoretinal microsurgery at previously inaccessible ocular sites. Many invasive excision and incision based diagnostic biopsy, prostrate, and nephrolgical procedures can be performed minimally or almost noninvasively due to recent advancements in microrobotic technology. Advances in biohybrid microrobot systems are pushing microrobotic systems even smaller, using biological cells as on-board microactuators and microsensors using the chemical energy. Such microrobotic systems could be used for local targeted delivery of imaging contrast agents, drugs, genes, and mRNA, minimally invasive surgery, and cell micromanipulation in the near future.

Miniaturized soft bio-hybrid robotics: a step forward into healthcare applications.

Soft robotics is an emerging discipline that employs soft flexible materials such as fluids, gels and elastomers in order to enhance the use of robotics in healthcare applications. Compared to their rigid counterparts, soft robotic systems have flexible and rheological properties that are closely related to biological systems, thus allowing the development of adaptive and flexible interactions with complex dynamic environments. With new technologies arising in bioengineering, the integration of living cells into soft robotic systems offers the possibility of accomplishing multiple complex functions such as sensing and actuating upon external stimuli. These emerging bio-hybrid systems are showing promising outcomes and opening up new avenues in the field of soft robotics for applications in healthcare and other fields.