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New acoustic-tweezer design allows for 3D bioprinting

Illustration of a particle (red sphere) trapped by the 3D trapping node created by two superimposed, orthogonal (at right angles), standing surface acoustic waves and induced acoustic streaming (credit: Carnegie Mellon University)

A team of researchers at three universities has developed a way to use “acoustic tweezers” (which use ultrasonic surface acoustic waves, or SAWs, to trap and manipulate micrometer-scale particles and biological cells — see “Acoustic tweezers manipulate cellular-scale objects with ultrasound“) to non-invasively pick up and move single cells in three mutually orthogonal axes of motion (three dimensions).

The new 3D acoustic tweezers can pick up single cells or entire cell assemblies and deliver them to desired locations to create 2D and 3D cell patterns, or print* the cells into complex shapes — a promising new method for “3D bioprinting” in biological tissues, the researchers say in an open-access paper in  the Proceedings of the National Academy of Sciences (PNAS).

The new method, developed by researchers at Carnegie Mellon University, Pennsylvania State University and MIT, offers the potential to accurately print 3D multicellular architectures for applications in biomanufacturing, tissue engineering, regenerative medicine, neuroscience, and cancer metastasis research.

Multicellular structures within living things are complex and delicate, which makes recreating or repairing these structures a daunting task. For example, the human heart contains more than 2 billion muscle cells. Each of these cells must properly interact with one another and with their environment to ensure that the heart functions properly. If those cells aren’t placed correctly, or are damaged, it could potentially result in any of a variety of heart conditions.

A 3D trapping node

Illustration of planar surface acoustic wave generators used to generate 3D nodes surrounding a microfluidic experimental area (center). The inset indicates a single particle within a “3-D trapping node” that is independently manipulated along x, y or z axes. (credit: Carnegie Mellon University)

Researchers have been using a combination of approaches for recreating the complex, multicellular architecture of biological tissue and to separate, align, pattern and transport single cells, and the approaches are renowned for their ability to gently manipulate cells without causing any cellular damage.

But they have yet to develop a single method that has the high level of precision, versatility, multiple dimensionality, and single cell resolution needed to form complex multicellular structures while maintaining cell viability, integrity and function and without the need for invasive contact, tagging, or biochemical labeling in regenerative medicine, neuroscience, tissue engineering, bio-manufacturing, and cancer metastasis.

The new microfluidic device developed by the team has now allowed the researchers to manipulate where the waves would meet along each of the three axes. At these meeting points, the waves formed a 3D trapping node that captured individual cells. The researchers could then further manipulate the acoustic waves to move and place cells.

Printing living cells with 3D acoustic tweezers. (A) Single-cell printing. After previously deposited 3T3 (mouse embryonic fibroblast) cells were attached to the substrate, another single cell was picked up, transported, and dropped at the desired location on the substrate (indicated by red arrow) or on top of another cell (indicated by blue arrow). The single cell adhered and spread along the surface. (B) Cell culture patterns arranged to form the characters “3,” “D,” “A,” and “T” by printing single HeLa S3 cells (a human immortal cell line) using 3D acoustic tweezers. (Scale bar: 20 micrometers) (credit: Feng Guo et al./PNAS)

To demonstrate how their acoustic tweezers technique could be used for live cell printing, the researchers used the microfluidic device to pick up cells and deposit them in a pre-selected pattern, with an elegant level of control over cell spacing and geometry. That suggested that the device has the potential to effectively create 3D tissue-like structures, including those with complex geometries.

* Typically, cells are printed in a substrate or matrix to form tissue for later implantation (see “Robust ‘spider silk’ matrix guides cardiac tissue regeneration“).

Abstract of Three-dimensional manipulation of single cells using surface acoustic waves

The ability of surface acoustic waves to trap and manipulate micrometer-scale particles and biological cells has led to many applications involving “acoustic tweezers” in biology, chemistry, engineering, and medicine. Here, we present 3D acoustic tweezers, which use surface acoustic waves to create 3D trapping nodes for the capture and manipulation of microparticles and cells along three mutually orthogonal axes. In this method, we use standing-wave phase shifts to move particles or cells in-plane, whereas the amplitude of acoustic vibrations is used to control particle motion along an orthogonal plane. We demonstrate, through controlled experiments guided by simulations, how acoustic vibrations result in micromanipulations in a microfluidic chamber by invoking physical principles that underlie the formation and regulation of complex, volumetric trapping nodes of particles and biological cells. We further show how 3D acoustic tweezers can be used to pick up, translate, and print single cells and cell assemblies to create 2D and 3D structures in a precise, noninvasive, label-free, and contact-free manner.

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Mechanotherapy may replace drug and cellular therapies for injured muscle tissue

These microscopic images reveal the dramatic effect that a novel mechanotherapy has on muscle regeneration of injured tissue over a period of two weeks: no treatment is pictured (left), vs. direct mechanical stimulation of the muscle (right), resulting in two-and-a-half-fold improvement in muscle regeneration, reduced tissue scarring and fibrosis, and a visible increase in the density of muscle cells. (credit: Wyss Institute at Harvard University)

Engineers and biomedical scientists at the Wyss Institute for Biologically Inspired Engineering at Harvard University and the Harvard School of Engineering and Applied Sciences have developed a promising new approach for repairing severely damaged skeletal muscles: direct mechanical stimulation. It may be appropriate for major injuries commonly caused by motor vehicle accidents, other traumas, or nerve damage, which can lead to extensive scarring, fibrous tissue, and loss of muscle function.

The new method could one day promote skeletal muscle regeneration and replace or enhance drug- and cell-based regenerative treatments. The finding was published on January 25 in the journal Proceedings of the National Academy of Sciences.

Two mechanotherapy methods of generating cyclic mechanical compressions (credit: Christine A. Cezar et al./PNAS)

The team investigated  two potential mechanotherapies for a two-week period, using mice with muscle injury and hind limb ischemia:

  • Ferrogel implant. They implanted a biocompatible magnetized gel called a “biphasic ferrogel,” located in direct contact with the damaged tissue. They then used external magnetic pulses to apply cyclic stimulation to the muscle.
  • Pressurized cuff. They fitted a soft robotic, non-invasive pressurized cuff over the injured leg and used pulses of air to cyclically massage the hind leg.

Finding: mechanical stimulation of muscle alone can foster regeneration 

To their surprise, the researchers discovered that both cyclic mechanical stimulation — provided by either magnetized gel or robotic cuff — resulted in a two-and-a-half-fold improvement in muscle regeneration and reduced tissue scarring over the course of two weeks (compared to no-treatment controls), ultimately leading to an improvement in muscle function.

That suggested that the use of non-invasive pressurized cuffs or devices alone could one day help heal patients suffering from severe muscle injuries — no invasive procedure required. The direct stimulation of muscle tissue increases the transport of oxygen, nutrients, fluids and waste removal from the site of the injury, which are all vital components of muscle health and repair, according to the reseachers.

Replacing drugs and cells

“Until now, most approaches to muscle regeneration have been biologic, relying on the use of drugs or cells,” said Christine Cezar, Ph.D., lead author on the study, who completed her doctoral research at the Wyss Institute and Harvard SEAS. “Our finding that mechanical stimulation alone is enough to enhance muscle repair could open the door to new non-biologic therapies, or even combinatorial therapies that employ both mechanical and biological interventions to treat severely damaged skeletal muscles.”

According to Harvard bioengineer David Mooney, Ph.D., senior author on the new study, one of the most exciting aspects of this research is that its translation to the clinic in the form of a stimulatory device could be relatively rapid as compared to drug or cell therapies. The team plans to explore how the findings can make the jump from the laboratory to the clinic.*

“Chemistry tends to dominate the way we think about medicine, but it has become clear that physical and mechanical factors play very critical roles in regulating biology,” said Mooney, a Wyss Institute Core Faculty member and the Robert P. Pinkas Family Professor of Bioengineering at the Harvard John A. Paulson School of Engineering and Applied Sciences (SEAS). “The results of our new study demonstrate how direct physical and mechanical intervention can impact biological processes and can potentially be exploited to improve clinical outcomes.”

Down the road, the principle of using mechanical stimulation to enhance regeneration or reduce formation of scarring or fibrosis could also be applied to a wide range of medical devices that interface mechanical components with body tissues, the researchers say. Currently, such medical devices are often plagued by the formation of thickened tissue capsules that form at the intersection of machine and man.

DIY muscle treatment?

The finding raises a question: what about testing conventional post-operative external pneumatic compression devices (such as an inflatable cuff or bootie that’s inflated and deflated to apply intermittent pressure) in clinic or at home (with physician oversight)? An improved device could have doctor- or patient-adjustable pressure settings for various levels of injury or various stages of healing. And what about deep massage**, which has the advantage of patient feedback on excessive pain (indicating possible tissue damage)?

* The multi-disciplinary team spanning the Wyss Institute’s Programmable Nanomaterials and Bioinspired Robotics platforms was led by Mooney and also included soft roboticist Conor Walsh, Ph.D., who is a Wyss Core Faculty member, Associate Professor of Mechanical and Biomedical Engineering at Harvard SEAS and Founder of the Harvard Biodesign Lab, and biomechanical engineer Georg Duda, Ph.D., who is a Wyss Associate Core Faculty member, Vice-Director of the Berlin-Brandenburg Center for Regenerative Therapies (BCRT) and the Director of the Julius Wolff Institute for Biomechanics and Musculoskeletal Regeneration at Charité-Universitätsmedizin Berlin.

** In the PNAS paper, the authors note that “a role for massage therapy as a potent immunomodulator after exercise-induced muscle damage in humans has been reported” and that there is some evidence that massage therapy or physical manipulation may promote recovery by increasing blood flow.

Abstract of Biologic-free mechanically induced muscle regeneration

Severe skeletal muscle injuries are common and can lead to extensive fibrosis, scarring, and loss of function. Clinically, no therapeutic intervention exists that allows for a full functional restoration. As a result, both drug and cellular therapies are being widely investigated for treatment of muscle injury. Because muscle is known to respond to mechanical loading, we investigated instead whether a material system capable of massage-like compressions could promote regeneration. Magnetic actuation of biphasic ferrogel scaffolds implanted at the site of muscle injury resulted in uniform cyclic compressions that led to reduced fibrous capsule formation around the implant, as well as reduced fibrosis and inflammation in the injured muscle. In contrast, no significant effect of ferrogel actuation on muscle vascularization or perfusion was found. Strikingly, ferrogel-driven mechanical compressions led to enhanced muscle regeneration and a ∼threefold increase in maximum contractile force of the treated muscle at 2 wk compared with no-treatment controls. Although this study focuses on the repair of severely injured skeletal muscle, magnetically stimulated bioagent-free ferrogels may find broad utility in the field of regenerative medicine.

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A new technique for super-resolution digital microscopy

The image sensor of the wavelength scanning super-resolution apparatus collects a “stack” of images of the sample (credit: Ozcan Lab)

Researchers from the California NanoSystems Institute at UCLA have created a new technique using lens-free holograms that greatly enhances digital microscopy images, which are sometimes blurry and pixelated.

The new technique, called “wavelength scanning pixel super-resolution,” uses a device that captures a stack of digital images of the same specimen, each with a slightly different wavelength of light. Then, researchers apply a newly devised algorithm that divides the pixels in each captured image into a number of smaller pixels, resulting in a much higher-resolution digital image of the specimen.

The research team was led by Aydogan Ozcan, Chancellor’s Professor of Electrical Engineering and Bioengineering at the UCLA Henry Samueli School of Engineering and Applied Science. The study appears in an open-access paper in the journal Light: Science and Applications, published by the Nature Publishing Group.

Raw data is transformed into a super-resolution image (credit: Ozcan Lab)

Faster, more-accessible diagnosis

“These results mean we can see and inspect large samples with finer details at the sub-micron [nanoscale] level,” Ozcan said. “We have applied this method to lens-based conventional microscopes, as well as our lensless on-chip microscopy systems that create microscopic images using holograms, and it works across all these platforms.”

The benefits of this new method are wide-ranging, but especially significant in pathology, where rapid microscopic imaging of large numbers of tissue or blood cells is key to diagnosing diseases such as cancer. The specimens used in the study were blood samples, used to screen for various diseases, and Papanicolaou tests, which are used to screen for cervical cancer.

Ozcan said that wavelength scanning super-resolution works on both colorless and dye-stained samples. The entire apparatus fits on a desktop, so its size and convenience could be of great benefit to doctors and scientists using microscopes in resource-limited settings such as clinics in developing countries.

The research was supported by the Presidential Early Career Award for Scientists and Engineers, the Army Research Office, the National Science Foundation, the Office of Naval Research and the Howard Hughes Medical Institute.

Abstract of Pixel super-resolution using wavelength scanning

Undersampling and pixelation affect a number of imaging systems, limiting the resolution of the acquired images, which becomes particularly significant for wide-field microscopy applications. Various super-resolution techniques have been implemented to mitigate this resolution loss by utilizing sub-pixel displacements in the imaging system, achieved, for example, by shifting the illumination source, the sensor-array and/or the sample, followed by digital synthesis of a smaller effective pixel by merging these subpixel-shifted low-resolution images. Herein, we introduce a new pixel super-resolution method that is based on wavelength scanning and demonstrate that as an alternative to physical shifting/displacements, wavelength diversity can be used to boost the resolution of a wide-field imaging system and significantly increase its space-bandwidth product. We confirmed the effectiveness of this new technique by improving the resolution of lens-free as well as lens-based microscopy systems and developed an iterative algorithm to generate high-resolution reconstructions of a specimen using undersampled diffraction patterns recorded at a few wavelengths covering a narrow spectrum (10-30 nm). When combined with a synthetic-aperture-based diffraction imaging technique, this wavelength-scanning super-resolution approach can achieve a half-pitch resolution of 250 nm, corresponding to a numerical aperture of approximately 1.0, across a large field of view (>20 mm2 ). We also demonstrated the effectiveness of this approach by imaging various biological samples, including blood and Papanicolaou smears. Compared with displacement-based super-resolution techniques, wavelength scanning brings uniform resolution improvement in all directions across a sensor array and requires significantly fewer measurements. This technique would broadly benefit wide-field imaging applications that demand larger space-bandwidth products.

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How cancer cells form tumors by reaching out with ‘cables’ and grabbing cells


University of Iowa | Cancer cells’ motion and accretion into tumors

Two University of Iowa studies have recorded the movements of cancerous human breast tissue cells in real time and in 3D — the first time cancer cells’ motion and accretion into tumors has been continuously tracked, the researchers believe.

The team discovered that cancerous cells, moving at move at 92 micrometers per hour (about twice the speed of healthy cells), actively recruit healthy cells into tumors by extending a kind of cable to grab their neighbors — both cancerous and healthy — and reel them in. Surprisingly, as little as five percent of cancerous cells are needed to form the tumors, a ratio previously unknown.

“It’s not like things sticking to each other,” said David Soll, biology professor at the UI and corresponding author on the open-access paper, published in the American Journal of Cancer Research. “It’s that these cells go out and actively recruit. It’s complicated stuff, and it’s not passive. No one had a clue that there were specialized cells in this process, and that it’s a small number that pulls all the rest in.”

The findings could lead to a more precise identification of tumorigenic cells (those that form tumors) and to testing which antibodies would be best equipped to eliminate them.*

How cancer cells “know” what to do

The question is: how do these cells know what to do. Soll hypothesizes they’re reaching back to a primitive past, when these cells were programmed to form embryos. If true, perhaps the cancerous cells — masquerading as embryo-forming cells — recruit other cells to make tissue that then forms the layered, self-sustaining architecture needed for a tumor to form and thrive. “It’s as if it’s building its own defenses against the body’s efforts to defeat them.”

University of Iowa researchers have documented how cancerous tumors form by tracking in real time the movement of individual cells in 3-D. They report that just 5 percent of cancer cells are needed to form tumors, a ratio that heretofore had been unknown. (credit: Soll Laboratory)

In the AJCR paper, the researchers found support for their previous observation that tumorigenic cell lines and fresh tumor cells possess the unique capacity to form tumors by the active formation of cellular cables.

The finding lends more weight to the idea that tumors are created concurrently, in multiple locations, by individual clusters of cells that employ the cancer-cell cables to draw in more cells and enlarge themselves. Some have argued that tumors come about more by cellular changes within the masses, known as the “cancer stem cell theory.”

The Developmental Studies Hybridoma Bank funded the study.

* Soll’s Monoclonal Antibody Research Institute and the Developmental Studies Hybridoma Bank, created by the National Institutes of Health as a national resource, directed by Soll and housed at the UI, together contain one of the world’s largest collections of antibodies that could be used for the anti-cancer testing, based on the new findings.

Abstract of Mediated coalescence: a possible mechanism for tumor cellular heterogeneity

Recently, we demonstrated that tumorigenic cell lines and fresh tumor cells seeded in a 3D Matrigel model, first grow as clonal islands (primary aggregates), then coalesce through the formation and contraction of cellular cables. Non-tumorigenic cell lines and cells from normal tissue form clonal islands, but do not form cables or coalesce. Here we show that as little as 5% tumorigenic cells will actively mediate coalescence between primary aggregates of majority non-tumorigenic or non-cancerous cells, by forming cellular cables between them. We suggest that this newly discovered, specialized characteristic of tumorigenic cells may explain, at least in part, why tumors contain primarily non-tumorigenic cells.

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Google machine-learning system is first to defeat professional Go player

Go is played on a grid of black lines (usually 19×19). Game pieces, called stones, are played on the line intersections. (credit: Goban1/Wikipedia)

A deep-learning computer system called AlphaGo created by Google’s DeepMind team has defeated reigning three-time European Go champion Fan Hui 5 games to 0 — the first time a computer program has ever beaten a professional Go player, reports Google Research blog today (Jan. 27) — a feat previously thought to be at least a decade away.

“AlphaGo uses general machine-learning techniques to allow it to improve itself, just by watching and playing games,” according to David Silver and Demis Hassabis of Google DeepMind. Using a vast collection of more than 30 million Go moves from expert players, DeepMind researchers trained their system to play Go on its own.

To achieve that, AlphaGo, as described in a paper in Nature today, combines a state-of-the-art tree search with two deep neural networks, each containsing many layers with millions of neuron-like connections needed to deal with Go’s vast search space — more than a googol (10100) times larger than chess (a number greater than there are atoms in the universe).

“We first trained [one of the two networks] on 30 million moves from games played by human experts, until it could predict the human move 57% of the time …. But our goal is to beat the best human players, not just mimic them, Silver and Hassabis said. “To do this, AlphaGo learned to discover new strategies for itself, by playing thousands of games between its neural networks, and gradually improving them using a trial-and-error process known as reinforcement learning.”

This figure from the Nature article shows the Elo rating (a 230 point gap corresponds to a 79% probability of winning) and approximate rank of AlphaGo (both single machine and distributed versions), the European champion Fan Hui (a professional 2-dan), and the strongest other Go programs, evaluated over thousands of games. Pale pink bars show the performance of other programs when given a four move headstart. (credit: David Silver et al./Nature)

AlphaGo’s next challenge will be to play the top Go player in the world over the last decade, Lee Sedol. The match will take place this March in Seoul, South Korea.

“While games are the perfect platform for developing and testing AI algorithms quickly and efficiently, ultimately we want to apply these techniques to important real-world problems,” the researchers say. “Because the methods we have used are general purpose, our hope is that one day they could be extended to help us address some of society’s toughest and most pressing problems, from climate modelling to complex disease analysis.”


Google DeepMind | Ground-breaking AlphaGo masters the game of Go

Abstract of Mastering the game of Go with deep neural networks and tree search

The game of Go has long been viewed as the most challenging of classic games for artificial intelligence owing to its enormous search space and the difficulty of evaluating board positions and moves. Here we introduce a new approach to computer Go that uses ‘value networks’ to evaluate board positions and ‘policy networks’ to select moves. These deep neural networks are trained by a novel combination of supervised learning from human expert games, and reinforcement learning from games of self-play. Without any lookahead search, the neural networks play Go at the level of state-of-the-art Monte Carlo tree search programs that simulate thousands of random games of self-play. We also introduce a new search algorithm that combines Monte Carlo simulation with value and policy networks. Using this search algorithm, our program AlphaGo achieved a 99.8% winning rate against other Go programs, and defeated the human European Go champion by 5 games to 0. This is the first time that a computer program has defeated a human professional player in the full-sized game of Go, a feat previously thought to be at least a decade away.

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A flexible, transparent pressure sensor

Pressure sensors wrap around and conform to the shape of the fingers while still accurately measuring pressure distribution. (credit: 2016 Someya Laboratory)

Doctors may one day be able to physically screen for breast cancer using pressure-sensitive rubber gloves to detect tumors, thanks to a transparent, bendable, and sensitive pressure sensor newly developed by Japanese and American teams.

Conventional pressure sensors can’t measure pressure changes accurately once they are twisted or wrinkled, making them unsuitable for use on complex and moving surfaces, and they can’t be miniaturized below 100 micrometers (0.1 millimeters) thickness because of limitations in current production methods.

To address these issues, an international team of researchers led by Dr. Sungwon Lee and Professor Takao Someya of the University of Tokyo’s Graduate School of Engineeringhas developed a nanofiber-type pressure sensor made from carbon nanotubes and graphene that can measure pressure distribution of rounded surfaces such as an inflated balloon and maintain its sensing accuracy even when bent over a radius of 80 micrometers, equivalent to just twice the width of a human hair. The sensor is roughly 8 micrometers thick and can measure the pressure in 144 locations at once.

The device demonstrated in this study consists of organic transistors, electronic switches made from carbon and oxygen-based organic materials, and a pressure-sensitive nanofiber structure. Carbon nanotubes and graphene were added to an elastic polymer to create nanofibers with a diameter of 300 to 700 nanometers, which were then entangled with each other to form a transparent, thin and light porous structure.

The material may also have applications in improving the touch sensitivity in robots.

Abstract of A transparent bending-insensitive pressure sensor

Measuring small normal pressures is essential to accurately evaluate external stimuli in curvilinear and dynamic surfaces such as natural tissues. Usually, sensitive and spatially accurate pressure sensors are achieved through conformal contact with the surface; however, this also makes them sensitive to mechanical deformation (bending). Indeed, when a soft object is pressed by another soft object, the normal pressure cannot be measured independently from the mechanical stress. Here, we show a pressure sensor that measures only the normal pressure, even under extreme bending conditions. To reduce the bending sensitivity, we use composite nanofibres of carbon nanotubes and graphene. Our simulations show that these fibres change their relative alignment to accommodate bending deformation, thus reducing the strain in individual fibres. Pressure sensitivity is maintained down to a bending radius of 80 μm. To test the suitability of our sensor for soft robotics and medical applications, we fabricated an integrated sensor matrix that is only 2 μm thick. We show real-time (response time of ∼20 ms), large-area, normal pressure monitoring under different, complex bending conditions.

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A novel ’4D printing’ method inspired by plants

This series of images shows the transformation (top right) of a 4D-printed hydrogel composite structure (top left) after its submersion (bottom) in water (credit: Wyss Institute at Harvard University)

Harvard University scientists have evolved their microscale 3D printing technology to the fourth dimension, time. Inspired by natural structures like plants, which respond and change their form over time according to environmental stimuli, the team has designed 4D-printed hydrogel composite structures that change shape upon immersion in water.

The team is located at the Wyss Institute for Biologically Inspired Engineering at Harvard University and the Harvard John A. Paulson School of Engineering and Applied Sciences.

“This work represents an elegant advance in programmable materials assembly, made possible by a multidisciplinary approach,” said Jennifer Lewis, Sc.D., senior author of a new study reported on January 25 in a new  in Nature Materials. “We have now gone beyond integrating form and function to create transformable architectures.”

In nature, flowers and plants have tissue compositions and microstructures that result in dynamic morphologies (forms) that change according to their environments. Mimicking the variety of shape changes undergone by plant organs such as tendrils, leaves, and flowers in response to environmental stimuli like humidity and/or temperature, the 4D-printed hydrogel composites developed by Lewis and her team are programmed to contain precise, localized swelling behaviors.

Derived from wood

The trick: the hydrogel composites contain cellulose fibrils that are derived from wood and are similar to the microstructures that enable shape changes in plants.

By aligning cellulose fibrils during printing, the hydrogel composite ink is encoded with anisotropic swelling and stiffness, which can be patterned to produce intricate shape changes. The anisotropic (irregular) nature of the cellulose fibrils gives rise to varied directional properties that can be predicted and controlled. That’s why wood can be split easier along the grain rather than across it.

Likewise, when immersed in water, the hydrogel-cellulose fibril ink undergoes differential swelling behavior along and orthogonal to the printing path. Combined with a proprietary mathematical model developed by the team that predicts how a 4D object must be printed to achieve prescribed transformable shapes, the new method opens up many new and exciting potential applications for 4D printing technology including smart textiles, soft electronics, biomedical devices, and tissue engineering.

The composite ink that the team uses flows like liquid through the printhead, yet rapidly solidifies once printed. A variety of hydrogel materials can be used interchangeably resulting in different stimuli-responsive behaviors, while the cellulose fibrils can be replaced with other anisotropic fillers of choice, including conductive fillers.

Complex flower morphologies generated by biomimetic 4D printing. A flower demonstrating a range of morphologies inspired by a native orchid, the Dendrobium helix (courtesy of Ricardo Valentin). Based on the print path, this orchid architecture exhibits four different configurations: bending, twisting and ruffling corolla surrounding the central funnel-like domain (scale bars, 5 mm). (credit: A. Sydney Gladman et al./Nature Materials)

The mathematical model prescribes the printing pathways required to achieve the desired shape-transforming response. Specifically, it solves the “inverse problem” — the challenge of being able to predict what the printing toolpath must be to encode swelling behaviors toward achieving a specific desired target shape.

“It is wonderful to be able to design and realize, in an engineered structure, some of nature’s solutions,” said L. Mahadevan, Ph.D., a Wyss Core Faculty member as well as the Lola England de Valpine Professor of Applied Mathematics, Professor of Organismic and Evolutionary Biology, and Professor of Physics at Harvard University and Harvard SEAS, is a co-author on the study. “By solving the inverse problem, we are now able to reverse-engineer the problem and determine how to vary local inhomogeneity, i.e. the spacing between the printed ink filaments, and the anisotropy, i.e. the direction of these filaments, to control the spatiotemporal response of these shapeshifting sheets.”

Lewis is a Core Faculty member at the Wyss Institute for Biologically Inspired Engineering at Harvard University and the Hansjörg Wyss Professor of Biologically Inspired Engineering at the Harvard John A. Paulson School of Engineering and Applied Science (SEAS). Their team also includes co-author, Ralph Nuzzo, Ph.D., the G.L. Clark Professor of Chemistry at the University of Illinois at Urbana-Champaign.


Wyss Institute | 4D Printing: Shapeshifting Architectures

Abstract of Biomimetic 4D printing

Shape-morphing systems can be found in many areas, including smart textiles, autonomous robotics, biomedical devices, drug delivery and tissue engineering. The natural analogues of such systems are exemplified by nastic plant motions, where a variety of organs such as tendrils, bracts, leaves and flowers respond to environmental stimuli (such as humidity, light or touch) by varying internal turgor, which leads to dynamic conformations governed by the tissue composition and microstructural anisotropy of cell walls. Inspired by these botanical systems, we printed composite hydrogel architectures that are encoded with localized, anisotropic swelling behaviour controlled by the alignment of cellulose fibrils along prescribed four-dimensional printing pathways. When combined with a minimal theoretical framework that allows us to solve the inverse problem of designing the alignment patterns for prescribed target shapes, we can programmably fabricate plant-inspired architectures that change shape on immersion in water, yielding complex three-dimensional morphologies.

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How to make almost any shape out of a flat sheet of paper

Mahadevan and his team have characterized a fundamental origami fold, or tessellation, that could be used as a building block to create almost any three-dimensional shape, as seen above (credit: Mahadevan Lab/Harvard SEAS)

Harvard scientist L. Mahadevan and his team have devised a way to make virtually any shape out of a flat sheet of paper, using a fundamental origami or tessellation fold.

The folding pattern, known as the Miura-ori, is a periodic way to tile the plane using the simplest mountain-valley fold in origami. It was used as a decorative item in clothing at least as long ago as the 15th century. A folded Miura can be packed into a flat, compact shape and unfolded in one continuous motion, making it ideal for packing rigid structures like solar panels.  It also occurs in nature in a variety of situations, such as in insect wings and certain leaves.

Mahadevan suggests that this simple folding pattern could serve as a template for more complicated shapes, such as space-bound payloads, surgical stents that can be packed flat and pop-up into three-dimensional structures once inside the body, or dining room tables that can lean flat against the wall until they are ready to be used.

To explore the potential of the tessellation, the team developed an algorithm that can create certain shapes using the Miura-ori fold, repeated with small variations. Given the specifications of the target shape, the program lays out the folds needed to create the design, which can then be laser printed for folding.

The program takes into account several factors, including the stiffness of the folded material and the trade-off between the accuracy of the pattern and the effort associated with creating finer folds — an important characterization because, as of now, these shapes are all folded by hand.

This spiral folds rigidly from flat pattern through the target surface and onto the flat-folded plane (credit: Mahadevan Lab)

“Essentially, we would like to be able to tailor any shape by using an appropriate folding pattern,” said Mahadevan. “Starting with the basic mountain-valley fold, our algorithm determines how to vary it by gently tweaking it from one location to the other to make a vase, a hat, a saddle, or to stitch them together to make more and more complex structures.”

“The really exciting thing about this fold is it is completely scalable.You can do this with graphene, which is one atom thick, or you can do it on the architectural scale.”

Mahadevan is the Lola England de Valpine Professor of Applied Mathematics, Organismic and Evolutionary Biology, and Physics at the Harvard John A. Paulson School of Engineering and Applied Sciences (SEAS). He is also a core faculty member of the Wyss Institute for Biologically Inspired Engineering, and member of the Kavli Institute for Bionano Science and Technology, at Harvard University.

The research is published in Nature Materials.

Abstract of Programming curvature using origami tessellations

Origami describes rules for creating folded structures from patterns on a flat sheet, but does not prescribe how patterns can be designed to fit target shapes. Here, starting from the simplest periodic origami pattern that yields one-degree-of-freedom collapsible structures—we show that scale-independent elementary geometric constructions and constrained optimization algorithms can be used to determine spatially modulated patterns that yield approximations to given surfaces of constant or varying curvature. Paper models confirm the feasibility of our calculations. We also assess the difficulty of realizing these geometric structures by quantifying the energetic barrier that separates the metastable flat and folded states. Moreover, we characterize the trade-off between the accuracy to which the pattern conforms to the target surface, and the effort associated with creating finer folds. Our approach enables the tailoring of origami patterns to drape complex surfaces independent of absolute scale, as well as the quantification of the energetic and material cost of doing so.

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New handheld miniature microscope could ID cancer cells in doctor’s offices and operating rooms

University of Washington mechanical engineers and collaborators have developed a handheld microscope to help doctors and dentists distinguish between healthy and cancerous cells in an office setting or operating room. (credit: Dennis Wise/University of Washington)

A miniature handheld microscope being developed by University of Washington mechanical engineers could allow neurosurgeons to differentiate cancerous from normal brain tissue at cellular level in real time in the operating room and determine where to stop cutting.

The new technology is intended to solve a critical problem in brain surgery: to definitively distinguish between cancerous and normal brain cells, during an operation, neurosurgeons would have stop the operation and send tissue samples to a pathology lab — where they are typically frozen, sliced, stained, mounted on slides and investigated under a bulky microscope.

Developed in collaboration with Memorial Sloan Kettering Cancer Center, Stanford University and the Barrow Neurological Institute, the new microscope is outlined in an open-access paper published in January in the journal Biomedical Optics Express.

“Surgeons don’t have a very good way of knowing when they’re done cutting out a tumor,” said senior author Jonathan Liu, UW assistant professor of mechanical engineering. “They’re using their sense of sight, their sense of touch, and pre-operative images of the brain — and oftentimes it’s pretty subjective. “Being able to zoom and see at the cellular level during the surgery would really help them to accurately differentiate between tumor and normal tissues and improve patient outcomes.”

The handheld microscope, roughly the size of a pen, combines technologies in a novel way to deliver high-quality images at faster speeds than existing devices. Researchers expect to begin testing it as a cancer-screening tool in clinical settings next year.

It also has other uses in medicine and dentistry. For instance, dentists who find a suspicious-looking lesion in a patient’s mouth often wind up cutting it out and sending it to a lab to be biopsied for oral cancer. Most come back benign.

That process subjects patients to an invasive procedure and overburdens pathology labs. A miniature microscope with high enough resolution to detect changes at a cellular level could be used in dental or dermatological clinics to better assess which lesions or moles are normal and which ones need to be biopsied.

Key technologies: dual-axis confocal microscopy and line scanning

The real-time microscope images (left) illuminate details in mouse tissues similar to the images (right) produced during an expensive, multi-day process at a clinical pathology lab. (credit: University of Washington)

“The microscope technologies that have been developed over the last couple of decades are expensive and still pretty large, about the size of a hair dryer or a small dental x-ray machine,” said co-author Milind Rajadhyaksha, associate faculty member in the dermatology service at the Memorial Sloan Kettering Cancer Center in New York City. “So there’s a need for creating much more miniaturized microscopes.”

Making microscopes smaller, however, usually requires sacrificing some aspect of image quality or performance such as resolution, field of view, depth, imaging contrast or processing speed.

“We feel like this device does one of the best jobs ever — compared to existing commercial devices and previous research devices — of balancing all those tradeoffs,” said Liu.

The miniature microscope uses an innovative approach called “dual-axis confocal microscopy” to illuminate and more clearly see through opaque tissue. It can capture details up to a half millimeter beneath the tissue surface, where some types of cancerous cells originate.

In the video below, for instance, researchers produced images of fluorescent blood vessels in a mouse ear at various depths ranging from 0.075 to 0.125 millimeters deep.

“Trying to see beneath the surface of tissue is like trying to drive in a thick fog with your high beams on – you really can’t see much in front of you,” Liu said. “But there are tricks we can play to see more deeply into the fog, like a fog light that illuminates from a different angle and reduces the glare.”

The microscope also employs a technique called line scanning to speed up the image-collection process. It uses micro-electrical-mechanical (MEMS) mirrors to direct an optical beam that scans the tissue, line by line, and quickly builds an image.

Imaging speed is particularly important for a handheld device, which has to contend with motion jitter from the human using it. If the imaging rate is too slow, the images will be blurry.

In the paper, the researchers demonstrate that the miniature microscope has sufficient resolution to see subcellular details. Images taken of mouse tissues compare well with those produced from a multi-day process at a clinical pathology lab — the gold standard for identifying cancerous cells in tissues.

The researchers hope that after testing the microscope’s performance as a cancer-screening tool, it can be introduced into surgeries or other clinical procedures within the next 2 to 4 years.

“For brain tumor surgery, there are often cells left behind that are invisible to the neurosurgeon. This device will really be the first to let you identify these cells during the operation and determine exactly how much further you can reduce this residual,” said project collaborator Nader Sanai, professor of neurosurgery at the Barrow Neurological Institute in Phoenix. “That’s not possible to do today.”

The research was funded by the National Institutes of Health through its National Institute of Dental and Craniofacial Research and National Cancer Institute.


University of Washington | Miniature dual-axis confocal microscope imaging

Abstract of Miniature in vivo MEMS-based line-scanned dual-axis confocal microscope for point-of-care pathology

There is a need for miniature optical-sectioning microscopes to enable in vivointerrogation of tissues as a real-time and noninvasive alternative to gold-standard histopathology. Such devices could have a transformative impact for the early detection of cancer as well as for guiding tumor-resection procedures. Miniature confocal microscopes have been developed by various researchers and corporations to enable optical sectioning of highly scattering tissues, all of which have necessitated various trade-offs in size, speed, depth selectivity, field of view, resolution, image contrast, and sensitivity. In this study, a miniature line-scanned (LS) dual-axis confocal (DAC) microscope, with a 12-mm diameter distal tip, has been developed for clinical point-of-care pathology. The dual-axis architecture has demonstrated an advantage over the conventional single-axis confocal configuration for reducing background noise from out-of-focus and multiply scattered light. The use of line scanning enables fast frame rates (16 frames/sec is demonstrated here, but faster rates are possible), which mitigates motion artifacts of a hand-held device during clinical use. We have developed a method to actively align the illumination and collection beams in a DAC microscope through the use of a pair of rotatable alignment mirrors. Incorporation of a custom objective lens, with a small form factor for in vivo clinical use, enables our device to achieve an optical-sectioning thickness and lateral resolution of 2.0 and 1.1 microns respectively. Validation measurements with reflective targets, as well as in vivo and ex vivo images of tissues, demonstrate the clinical potential of this high-speed optical-sectioning microscopy device.

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Planet Nine from outer space

This artistic rendering shows the distant view from Planet Nine back towards the sun. The planet is thought to be gaseous, similar to Uranus and Neptune. Hypothetical lightning lights up the night side. (credit: Caltech/R. Hurt (IPAC))

Caltech researchers have found evidence of a giant planet tracing a bizarre, highly elongated orbit in the outer solar system that the researchers have nicknamed Planet Nine.

It has a mass about ten times that of Earth and orbits about 20 times farther from the sun on average than does Neptune (which orbits the sun at an average distance of 2.8 billion miles). In fact, it would take this new planet between 10,000 and 20,000 years to make just one full orbit around the sun.

The researchers, Konstantin Batygin and Mike Brown (best known for the significant role he played in the demotion of Pluto from a planet to a dwarf planet), discovered the planet’s existence through mathematical modeling and computer simulations but have not yet observed the object directly.

“This would be a real ninth planet,” says Brown, the Richard and Barbara Rosenberg Professor of Planetary Astronomy.

Brown notes that the putative ninth planet—at 5,000 times the mass of Pluto—is sufficiently large that there should be no debate about whether it is a true planet. Unlike the class of smaller objects now known as dwarf planets, Planet Nine gravitationally dominates its neighborhood of the solar system. In fact, it dominates a region larger than any of the other known planets—a fact that Brown says makes it “the most planet-y of the planets in the whole solar system.”

Batygin and Brown describe their work in an open-access paper in the current issue of the Astronomical Journal and show how Planet Nine helps explain a number of mysterious features of the field of icy objects and debris beyond Neptune known as the Kuiper Belt. “For the first time in over 150 years, there is solid evidence that the solar system’s planetary census is incomplete,” said Batygin.

Planet Nine’s existence also helps explain the alignment of distant Kuiper Belt objects and the mysterious orbits that two of them trace.

The six most distant known objects in the solar system with orbits exclusively beyond Neptune (magenta) all mysteriously line up in a single direction. Also, when viewed in three dimensions, they tilt nearly identically away from the plane of the solar system. Batygin and Brown show that a planet with 10 times the mass of the earth in a distant eccentric orbit anti-aligned with the other six objects (orange) is required to maintain this configuration. (credit: Caltech/R. Hurt (IPAC); [Diagram created using WorldWide Telescope.])

So where did Planet Nine come from and how did it end up in the outer solar system? Scientists have long believed that the early solar system began with four planetary cores that went on to grab all of the gas around them, forming the four gas planets — Jupiter, Saturn, Uranus, and Neptune. Over time, collisions and ejections shaped them and moved them out to their present locations.

“But there is no reason that there could not have been five cores, rather than four,” says Brown. Planet Nine could represent that fifth core, and if it got too close to Jupiter or Saturn, it could have been ejected into its distant, eccentric orbit.

A predicted consequence of Planet Nine is that a second set of confined objects should also exist. These objects are forced into positions at right angles to Planet Nine and into orbits that are perpendicular to the plane of the solar system. Five known objects (blue) fit this prediction precisely. (credit: Caltech/R. Hurt (IPAC) [Diagram was created using WorldWide Telescope.])

Replacing Pluto

Only the planet’s rough orbit is known, not the precise location of the planet on that elliptical path. If the planet happens to be close to its perihelion, Brown says, astronomers should be able to spot it in images captured by previous surveys. If it is in the most distant part of its orbit, the world’s largest telescopes — such as the twin 10-meter telescopes at the W. M. Keck Observatory and the Subaru Telescope, all on Mauna Kea in Hawaii — will be needed to see it. If, however, Planet Nine is now located anywhere in between, many telescopes have a shot at finding it.

“One of the most startling discoveries about other planetary systems has been that the most common type of planet out there has a mass between that of Earth and that of Neptune,” says Batygin. “Until now, we’ve thought that the solar system was lacking in this most common type of planet. Maybe we’re more normal after all.”

“All those people who are mad that Pluto is no longer a planet can be thrilled to know that there is a real planet out there still to be found,” Brown says. “Now we can go and find this planet and make the solar system have nine planets once again.”


Caltech AMT | Caltech’s Konstantin Batygin, an assistant professor of planetary science, and Mike Brown, the Richard and Barbara Rosenberg Professor of Planetary Astronomy, discuss new research that provides evidence of a giant planet tracing a bizarre, highly elongated orbit in the outer solar system.

Abstract of Evidence for a Distant Giant Planet in the Solar System

Recent analyses have shown that distant orbits within the scattered disk population of the Kuiper Belt exhibit an unexpected clustering in their respective arguments of perihelion. While several hypotheses have been put forward to explain this alignment, to date, a theoretical model that can successfully account for the observations remains elusive. In this work we show that the orbits of distant Kuiper Belt objects (KBOs) cluster not only in argument of perihelion, but also in physical space. We demonstrate that the perihelion positions and orbital planes of the objects are tightly confined and that such a clustering has only a probability of 0.007% to be due to chance, thus requiring a dynamical origin. We find that the observed orbital alignment can be maintained by a distant eccentric planet with mass gsim10 m⊕ whose orbit lies in approximately the same plane as those of the distant KBOs, but whose perihelion is 180° away from the perihelia of the minor bodies. In addition to accounting for the observed orbital alignment, the existence of such a planet naturally explains the presence of high-perihelion Sedna-like objects, as well as the known collection of high semimajor axis objects with inclinations between 60° and 150° whose origin was previously unclear. Continued analysis of both distant and highly inclined outer solar system objects provides the opportunity for testing our hypothesis as well as further constraining the orbital elements and mass of the distant planet.