Three-dimensional cell agglomerates are broadly useful in tissue engineering and drug testing. We report a well-free method to form large (1.4-mm) multicellular clusters using 100-MHz surface acoustic waves (SAW) without direct contact with the media or cells. A fluid couplant is used to transform the SAW into acoustic streaming in the cell-laden media held in a petri dish. The couplant transmits longitudinal sound waves, forming a Lamb wave in the petri dish that, in turn, produces longitudinal sound in the media. Due to recirculation, human embryonic kidney (HEK293) cells in the dish are carried to the center of the coupling location, forming a cluster in less than 10 min. A few minutes later, these clusters may then be translated and merged to form large agglomerations, and even repeatedly folded to produce a roughly spherical shape of over 1.4 mm in diameter for incubation-without damaging the existing intercellular bonds. Calcium ion signaling through these clusters and confocal images of multiprotein junctional complexes suggest a continuous tissue construct: intercellular communication. They may be formed at will, and the method is feasibly useful for formation of numerous agglomerates in a single petri dish.

5' AMP-activated protein kinase (AMPK) is a highly conserved serine-threonine kinase that regulates energy expenditure by activating catabolic metabolism and suppressing anabolic pathways to increase cellular energy levels. Therefore AMPK activators are considered to be drug targets for treatment of metabolic diseases such as diabetes mellitus. To identify novel AMPK activators, we screened xanthene derivatives. We determined that the AMPK activators 9H-xanthene-9-carboxylic acid {2,2,2-trichloro-1-[3-(3-nitro-phenyl)-thioureido]-ethyl}-amide (Xn) and 9H-xanthene-9-carboxylic acid {2,2,2-trichloro-1-[3-(3-cyano-phenyl)-thioureido]-ethyl}-amide (Xc) elevated glucose uptake in L6 myotubes by stimulating translocation of glucose transporter type 4 (GLUT4). Treatment with the chemical AMPK inhibitor compound C and infection with dominant-negative AMPKa2-virus inhibited AMPK phosphorylation and glucose uptake in myotubes induced by either Xn or Xc. Of the two major upstream kinases of AMPK, we found that Xn and Xc showed LKB1 dependency by knockdown of STK11, an ortholog of human LKB1. Single intravenous administration of Xn and Xc to high-fat diet-induced diabetic mice stimulated AMPK phosphorylation of skeletal muscle and improved glucose tolerance. Taken together, these results suggest that Xn and Xc regulate glucose homeostasis through LKB1-dependent AMPK activation and that the compounds are potential candidate drugs for the treatment of type 2 diabetes mellitus.

Ultrasound has been used to non-invasively manipulate neuronal functions in humans and other animals. However, this approach is limited as it has been challenging to target specific cells within the brain or body. Here, we identify human Transient Receptor Potential A1 (hsTRPA1) as a candidate that confers ultrasound sensitivity to mammalian cells. Ultrasound-evoked gating of hsTRPA1 specifically requires its N-terminal tip region and cholesterol interactions; and target cells with an intact actin cytoskeleton, revealing elements of the sonogenetic mechanism. Next, we use calcium imaging and electrophysiology to show that hsTRPA1 potentiates ultrasound-evoked responses in primary neurons. Furthermore, unilateral expression of hsTRPA1 in mouse layer V motor cortical neurons leads to c-fos expression and contralateral limb responses in response to ultrasound delivered through an intact skull. Collectively, we demonstrate that hsTRPA1-based sonogenetics can effectively manipulate neurons within the intact mammalian brain, a method that could be used across species.

Ultrasound has been used to manipulate cells in both humans and animal models. While intramembrane cavitation and lipid clustering have been suggested as likely mechanisms, they lack experimental evidence. Here, high-speed digital holographic microscopy (kiloHertz order) is used to visualize the cellular membrane dynamics. It is shown that neuronal and fibroblast membranes deflect about 150 nm upon ultrasound stimulation. Next, a biomechanical model that predicts changes in membrane voltage after ultrasound exposure is developed. Finally, the model predictions are validated using whole-cell patch clamp electrophysiology on primary neurons. Collectively, it is shown that ultrasound stimulation directly defects the neuronal membrane leading to a change in membrane voltage and subsequent depolarization. The model is consistent with existing data and provides a mechanism for both ultrasound-evoked neurostimulation and sonogenetic control.

Hydrogen production techniques based on solar-water splitting have emerged as carbon-free energy systems. Many researchers have developed highly efficient thin-film photoelectrochemical (PEC) devices made of low-cost and earth-abundant materials. However, solar water splitting systems suffer from short lifetimes due to catalyst instability that is attributed to both chemical dissolution and mechanical stress produced by hydrogen bubbles. A recent study found that the nanoporous hydrogel could prevent the structural degradation of the PEC devices. In this study, we investigate the protection mechanism of the hydrogel-based overlayer by engineering its porous structure using the cryogelation technique. Tests for cryogel overlayers with varied pore structures, such as disconnected micropores, interconnected micropores, and surface macropores, reveal that the hydrogen gas trapped in the cryogel protector reduce shear stress at the catalyst surface by providing bubble nucleation sites. The cryogelated overlayer effectively preserves the uniformly distributed platinum catalyst particles on the device surface for over 200 h. Our finding can help establish semi-permanent photoelectrochemical devices to realize a carbon-free society.

Collective cell migration plays a critical role in physiological and pathological processes such as development, wound healing, and metastasis. Numerous studies have demonstrated how various types of chemical, mechanical, and electrical cues dictate the collective migratory behaviors of cells. Although an acoustic cue can be advantageous because of its noninvasiveness and biocompatibility, cell migration in response to acoustic stimulation remains poorly understood. In this study, we developed a device that is able to apply surface acoustic waves to a cell culture substrate and investigated the effect of propagating acoustic waves on collective cell migration. The migration distance estimated at various wave intensities revealed that unidirectional cell migration was enhanced at a critical wave intensity and that it was suppressed as the intensity was further increased. The increased migration might be attributable to cell orientation alignment along the direction of the propagating wave, as characterized by nucleus shape. Thicker actin bundles indicative of a high traction force were observed in cells subjected to propagating acoustic waves at the critical intensity. Our device and technique can be useful for regulating cellular functions associated with cell migration.

Ultrasound has been shown to affect the function of both neurons and non-neuronal cells, but, the underlying molecular machinery has been poorly understood. Here, we show that at least two mechanosensitive proteins act together to generate C. elegans behavioral responses to ultrasound stimuli. We first show that these animals generate reversals in response to a single 10 msec pulse from a 2.25 MHz ultrasound transducer. Next, we show that the pore-forming subunit of the mechanosensitive channel TRP-4, and a DEG/ENaC/ASIC ion channel MEC-4, are both required for this ultrasound-evoked reversal response. Further, the trp-4;mec-4 double mutant shows a stronger behavioral deficit compared to either single mutant. Finally, overexpressing TRP-4 in specific chemosensory neurons can rescue the ultrasound-triggered behavioral deficit in the mec-4 null mutant, suggesting that both TRP-4 and MEC-4 act together in affecting behavior. Together, we demonstrate that multiple mechanosensitive proteins likely cooperate to transform ultrasound stimuli into behavioral changes.

How the Venus flytrap (Dionaea muscipula) evolved the remarkable ability to sense, capture, and digest animal prey for nutrients has long puzzled the scientific community.1 Recent genome and transcriptome sequencing studies have provided clues to the genes thought to play a role in these tasks.2,3,4,5 However, proving a causal link between these and any aspect of the plant's hunting behavior has been challenging due to the genetic intractability of this non-model organism. Here, we use CRISPR-Cas9 methods to generate targeted modifications in the Venus flytrap genome. The plant detects prey using touch-sensitive trigger hairs located on its bilobed leaves.6 Upon bending, these hairs convert mechanical touch signals into changes in the membrane potential of sensory cells, leading to rapid closure of the leaf lobes to ensnare the animal.7 Here, we generate mutations in trigger-hair-expressed MscS-like (MSL)-family mechanosensitive ion channel genes FLYCATCHER1 (FLYC1) and FLYCATCHER2 (FLYC2)5 and find that double-mutant plants have a reduced leaf-closing response to mechanical ultrasound stimulation. While we cannot exclude off-target effects of the CRISPR-Cas9 system, our genetic analysis is consistent with these and other functionally redundant mechanosensitive ion channels acting together to generate the sensory system necessary for prey detection.

The fabrication of functional tissues is essential for clinical applications such as disease treatment and drug discovery. Recent studies have revealed that the mechanical environments of tissues, determined by geometric cell patterns, material composition, or mechanical properties, play critical roles in ensuring proper tissue function. Here, we propose an acoustophoretic technique using surface acoustic waves to fabricate therapeutic vascular tissue containing a three-dimensional collateral distribution of vessels. Co-aligned human umbilical vein endothelial cells and human adipose stem cells that are arranged in a biodegradable catechol-conjugated hyaluronic acid hydrogel exhibit enhanced cell-cell contacts, gene expression, and secretion of angiogenic and anti-inflammatory paracrine factors. The therapeutic effects of the fabricated vessel constructs are demonstrated in experiments using an ischemia mouse model by exhibiting the remarkable recovery of damaged tissue. Our study can be referenced to fabricate various types of artificial tissues that mimic the original functions as well as structures.

The development of diagnostics and medical devices has historically been concentrated in high-income countries, despite a significant need to expand healthcare services to low- and middle-income countries (LMIC). Poor quality healthcare extends beyond LMIC to underserved communities in developed countries. This paper reviews diseases and conditions that have not received much attention in the past despite imposing a significant burden on healthcare systems in these circumstances. We review the underlying mechanism of action of these conditions and current technology in use for diagnosis or surgical intervention. We aim to identify areas for technological development and review policy considerations that will enable real-world adoption. Specifically, this review focuses on diseases prevalent in sub-Saharan Africa and south Asia: melioidosis, infant and maternal mortality, schistosomiasis, and heavy metal and pesticide poisoning. Our aim with this review is to identify problems facing the world that require the attention of the medical device community and provide recommendations for research directions for groups interested in this field.