In Vivo Porcine Model of Acute Iliocaval Deep Vein Thrombosis.

CLINICAL IMPACT: The study establishes a rapid, technically straightforward, and reproducible porcine large animal model for acute iliocaval deep vein thrombosis (DVT). The procedure can be performed with basic endovascular skillsets. With its procedural efficiency and consistency, the platform is promising for comparative in vivo testing of venous thrombectomy devices in a living host, and for future verification and validation studies to determine efficacy of novel thrombectomy devices relative to predicates.

Dynamic control of contractile resistance to iPSC-derived micro-heart muscle arrays.

Many types of cardiovascular disease are linked to the mechanical forces placed on the heart. However, our understanding of how mechanical forces exactly affect the cellular biology of the heart remains incomplete. In vitro models based on cardiomyocytes derived from human induced pluripotent stem cells (iPSC-CM) enable researchers to develop medium to high-throughput systems to study cardiac mechanobiology at the cellular level. Previous models have been developed to enable the study of mechanical forces, such as cardiac afterload. However, most of these models require exogenous extracellular matrix (ECM) to form cardiac tissues. Recently, a system was developed to simulate changes in afterload by grafting ECM-free micro-heart muscle arrays to elastomeric substrates of discrete stiffnesses. In the present study, we extended this system by combining the elastomer-grafted tissue arrays with a magnetorheological elastomeric substrate. This system allows iPSC-CM based micro-heart muscle arrays to experience dynamic changes in contractile resistance to mimic dynamically altered afterload. Acute changes in substrate stiffness led to acute changes in the calcium dynamics and contractile forces, illustrating the system's ability to dynamically elicit changes in tissue mechanics by dynamically changing contractile resistance.

Plasmon-Enhanced Digital Fluoroimmunoassay for Subfemtomolar Detection of Protein Biomarkers.

Rapid and accurate quantification of low-abundance protein biomarkers in biofluids can transform the diagnosis of a range of pathologies, including infectious diseases. Here, we harness ultrabright plasmonic fluors as "digital nanolabels" and demonstrate the detection and quantification of subfemtomolar concentrations of human IL-6 and SARS-CoV-2 alpha and variant proteins in clinical nasopharyngeal swab and saliva samples from COVID-19 patients. The resulting digital plasmonic fluor-linked immunosorbent assay (digital p-FLISA) enables detection of SARS-CoV-2 nucleocapsid protein, both in solution and in live virions. Digital p-FLISA outperforms the "gold standard" enzyme-linked immunosorbent assay (ELISA), having a nearly 7000-fold lower limit-of-detection, and outperforms a commercial antigen test, having over 5000-fold improvement in analytical sensitivity. Detection and quantification of very low concentrations of target proteins holds potential for early detection of pathological conditions, treatment monitoring, and personalized medicine.

Genetically Engineered Protein-Based Bioadhesives with Programmable Material Properties.

Silk-amyloid-mussel foot protein (SAM) hydrogels made from recombinant fusion proteins containing ß-amyloid peptide, spider silk domain, and mussel foot protein (Mfp) are attractive bioadhesives as they display a unique combination of tunability, biocompatibility, bioabsorbability, strong cohesion, and underwater adhesion to a wide range of biological surfaces. To design tunable SAM hydrogels for tailored surgical repair applications, an understanding of the relationships between protein sequence and hydrogel properties is imperative. Here, we fabricated SAM hydrogels using fusion proteins of varying lengths of silk-amyloid repeats and Mfps to characterize their structure and properties. We found that increasing silk-amyloid repeats enhanced the hydrogel's ß-sheet content (r = 0.74), leading to higher cohesive strength and toughness. Additionally, increasing the Mfp length beyond the half-length of the full Mfp sequence (1/2 Mfp) decreased the ß-sheet content (r = -0.47), but increased hydrogel surface adhesion. Among different variants, the hydrogel made of 16xKLV-2Mfp displayed a high ultimate strength of 3.0 ± 0.3 MPa, an ultimate strain of 664 ± 119%, and an attractive underwater adhesivity of 416 ± 20 kPa to porcine skin. Collectively, the sequence-structure-property relationships learned from this study will be useful to guide the design of future protein adhesives with tunable characteristics for tailored surgical applications.

Programmable and Reversible Integrin-Mediated Cell Adhesion Reveals Hysteresis in Actin Kinetics that Alters Subsequent Mechanotransduction.

Dynamically evolving adhesions between cells and extracellular matrix (ECM) transmit time-varying signals that control cytoskeletal dynamics and cell fate. Dynamic cell adhesion and ECM stiffness regulate cellular mechanosensing cooperatively, but it has not previously been possible to characterize their individual effects because of challenges with controlling these factors independently. Therefore, a DNA-driven molecular system is developed wherein the integrin-binding ligand RGD can be reversibly presented and removed to achieve cyclic cell attachment/detachment on substrates of defined stiffness. Using this culture system, it is discovered that cyclic adhesion accelerates F-actin kinetics and nuclear mechanosensing in human mesenchymal stem cells (hMSCs), with the result that hysteresis can completely change how hMSCs transduce ECM stiffness. Results are dramatically different from well-known results for mechanotransduction on static substrates, but are consistent with a mathematical model of F-actin fragments retaining structure following loss of integrin ligation and participating in subsequent repolymerization. These findings suggest that cyclic integrin-mediated adhesion alters the mechanosensing of ECM stiffness by hMSCs through transient, hysteretic memory that is stored in F-actin.

Virtual blebbistatin: A robust and rapid software approach to motion artifact removal in optical mapping of cardiomyocytes.

Fluorescent reporters of cardiac electrophysiology provide valuable information on heart cell and tissue function. However, motion artifacts caused by cardiac muscle contraction interfere with accurate measurement of fluorescence signals. Although drugs such as blebbistatin can be applied to stop cardiac tissue from contracting by uncoupling calcium-contraction, their usage prevents the study of excitation-contraction coupling and, as we show, impacts cellular structure. We therefore developed a robust method to remove motion computationally from images of contracting cardiac muscle and to map fluorescent reporters of cardiac electrophysiological activity onto images of undeformed tissue. When validated on cardiomyocytes derived from human induced pluripotent stem cells (iPSCs), in both monolayers and engineered tissues, the method enabled efficient and robust reduction of motion artifact. As with pharmacologic approaches using blebbistatin for motion removal, our algorithm improved the accuracy of optical mapping, as demonstrated by spatial maps of calcium transient decay. However, unlike pharmacologic motion removal, our computational approach allowed direct analysis of calcium-contraction coupling. Results revealed calcium-contraction coupling to be more uniform across cells within engineered tissues than across cells in monolayer culture. The algorithm shows promise as a robust and accurate tool for optical mapping studies of excitation-contraction coupling in heart tissue.

Stability of navigation in catheter-based endovascular procedures.

Endovascular procedures provide surgeons and other interventionalists with minimally invasive methods to treat vascular diseases by passing guidewires, catheters, sheaths and treatment devices into the vasculature to and navigate toward a treatment site. The efficiency of this navigation affects patient outcomes, but is frequently compromised by catheter "herniation", in which the catheter-guidewire system bulges out from the intended endovascular pathway so that the interventionalist can no longer advance it. Here, we showed herniation to be a bifurcation phenomenon that can be predicted and controlled using mechanical characterizations of catheter-guidewire systems and patientspecific clinical imaging. We demonstrated our approach in laboratory models and, retrospectively, in patients who underwent procedures involving transradial neurovascular procedures with an endovascular pathway from the wrist, up in the arm, around the aortic arch, and into the neurovasculature. Our analyses identified a mathematical navigation stability criterion that predicted herniation in all of these settings. Results show that herniation can be predicted through bifurcation analysis, and provide a framework for selecting catheter-guidewire systems to avoid herniation in specific patient anatomy.

Application of the Interagency and Modeling Analysis Group Model Verification Approach for Scientific Reproducibility in a Study of Biomineralization.

Model verification is a critical aspect of scientific accountability, transparency, and learning. Here, we demonstrate an application of a model verification approach for a molecular dynamics (MD) simulation, where the interactions between silica and silk protein were studied experimentally toward understanding biomineralization. Following the ten rules for credible modeling and simulation of biosciences as developed in Erdemir et al., the authors of the original paper collaborated with an external modeling group to verify the key findings of their original simulation model and to document this verification approach. The process resulted in successful replication of the key findings of the original model. Beyond verification, study of the model from a new perspective generated new insight into the basic assumptions. We discuss key learnings for how model validation processes can be improved more generally, specifically through improved documentation methods. We anticipate that this application of our protocol for model verification can be further replicated and improved to verify and validate other simulations.

Encoding and Storage of Information in Mechanical Metamaterials.

Information processing using material's own properties has gained increasing interest. Mechanical metamaterials, due to their diversity of deformation modes and wide design space, can be used to realize information processing, such as computing and storage. Here a mechanical metamaterial system is demonstrated for material-based encoding and storage of data through programmed reconfigurations of the metamaterial's structured building blocks. Sequential encoding and decoding are achieved in the three-dimensional (3D) printed pixelated mechanical metamaterial via kirigami-based "pixels" with programmable, temperature-dependent bistability. The mechanical metamaterial is demonstrated via a multistep deformation of encoding messages of texts and surfaces with arrays of binary data, and then decoding them by applying a predetermined stretching and heating regimen to sequentially retrieve layers of stored information and display them on its surface. This approach serves as a general framework to enable the encoding and storage of data with mechanical metamaterials.

Mechanobiology of cancer cell responsiveness to chemotherapy and immunotherapy: Mechanistic insights and biomaterial platforms.

Mechanical forces are central to how cancer treatments such as chemotherapeutics and immunotherapies interact with cells and tissues. At the simplest level, electrostatic forces underlie the binding events that are critical to therapeutic function. However, a growing body of literature points to mechanical factors that also affect whether a drug or an immune cell can reach a target, and to interactions between a cell and its environment affecting therapeutic efficacy. These factors affect cell processes ranging from cytoskeletal and extracellular matrix remodeling to transduction of signals by the nucleus to metastasis of cells. This review presents and critiques the state of the art of our understanding of how mechanobiology impacts drug and immunotherapy resistance and responsiveness, and of the in vitro systems that have been of value in the discovery of these effects.

Ultrasensitive lateral-flow assays via plasmonically active antibody-conjugated fluorescent nanoparticles.

Lateral-flow assays (LFAs) are rapid and inexpensive, yet they are nearly 1,000-fold less sensitive than laboratory-based tests. Here we show that plasmonically active antibody-conjugated fluorescent gold nanorods can make conventional LFAs ultrasensitive. With sample-to-answer times within 20 min, plasmonically enhanced LFAs read out via a standard benchtop fluorescence scanner attained about 30-fold improvements in dynamic range and in detection limits over 4-h-long gold-standard enzyme-linked immunosorbent assays, and achieved 95% clinical sensitivity and 100% specificity for antibodies in plasma and for antigens in nasopharyngeal swabs from individuals with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Comparable improvements in the assay's performance can also be achieved via an inexpensive portable scanner, as we show for the detection of interleukin-6 in human serum samples and of the nucleocapsid protein of SARS-CoV-2 in nasopharyngeal samples. Plasmonically enhanced LFAs outperform standard laboratory tests in sensitivity, speed, dynamic range, ease of use and cost, and may provide advantages in point-of-care diagnostics.

Nonlinear time-dependent mechanical behavior of mammalian collagen fibrils.

The viscoelastic mechanical behavior of collagenous tissues has been studied extensively at the macroscale, yet a thorough quantitative understanding of the time-dependent mechanics of the basic building blocks of tissues, the collagen fibrils, is still missing. In order to address this knowledge gap, stress relaxation and creep tests at various stress (5-35 MPa) and strain (5-20%) levels were performed with individual collagen fibrils (average diameter of fully hydrated fibrils: 253 ± 21 nm) in phosphate buffered saline (PBS). The experimental results showed that the time-dependent mechanical behavior of fully hydrated individual collagen fibrils reconstituted from Type I calf skin collagen, is described by strain-dependent stress relaxation and stress-dependent creep functions in both the heel-toe and the linear regimes of deformation in monotonic stress-strain curves. The adaptive quasilinear viscoelastic (QLV) model, originally developed to capture the nonlinear viscoelastic response of collagenous tissues, provided a very good description of the nonlinear stress relaxation and creep behavior of the collagen fibrils. On the other hand, the nonlinear superposition (NSP) model fitted well the creep but not the stress relaxation data. The time constants and rates extracted from the adaptive QLV and the NSP models, respectively, pointed to a faster rate for stress relaxation than creep. This nonlinear viscoelastic behavior of individual collagen fibrils agrees with prior studies of macroscale collagenous tissues, thus demonstrating consistent time-dependent behavior across length scales and tissue hierarchies. STATEMENT OF SIGNIFICANCE: Pure stress relaxation and creep experiments were conducted for the first time with fully hydrated individual collagen fibrils. It is shown that collagen nanofibrils have a nonlinear time-dependent behavior which agrees with prior studies on macroscale collagenous tissues, thus demonstrating consistent time-dependent behavior across length scales and tissue hierarchies. This new insight into the non-linear viscoelastic behavior of the building blocks of mammalian collagenous tissues may serve as the foundation for improved macroscale tissue models that capture the mechanical behavior across length scales.

Predicting YAP/TAZ nuclear translocation in response to ECM mechanosensing.

Cells translate mechanical cues from the extracellular matrix (ECM) into signaling that can affect the nucleus. One pathway by which such nuclear mechanotransduction occurs is a signaling axis that begins with integrin-ECM bonds and continues through a cascade of chemical reactions and structural changes that lead to nuclear translocation of YAP/TAZ. This signaling axis is self-reinforcing, with stiff ECM promoting integrin binding and thus facilitating polymerization and tension in the cytoskeletal contractile apparatus, which can compress nuclei, open nuclear pore channels, and enhance nuclear accumulation of YAP/TAZ. We previously developed a computational model of this mechanosensing axis for the linear elastic ECM by assuming that there is a linear relationship between the nucleocytoplasmic ratio of YAP/TAZ and nuclear flattening. Here, we extended our previous model to more general ECM behaviors (e.g., viscosity, viscoelasticity, and viscoplasticity) and included detailed YAP/TAZ translocation dynamics based on nuclear deformation. This model was predictive of diverse mechanosensing responses in a broad range of cells. Results support the hypothesis that diverse mechanosensing phenomena across many cell types arise from a simple, unified set of mechanosensing pathways.

An ECM-Mimicking, Injectable, Viscoelastic Hydrogel for Treatment of Brain Lesions.

Brain lesions can arise from traumatic brain injury, infection, and craniotomy. Although injectable hydrogels show promise for promoting healing of lesions and health of surrounding tissue, enabling cellular ingrowth and restoring neural tissue continue to be challenging. It is hypothesized that these challenges arise in part from the mismatch of composition, stiffness, and viscoelasticity between the hydrogel and the brain parenchyma, and this hypothesis is tested by developing and evaluating a self-healing hydrogel that not only mimics the composition, but also the stiffness and viscoelasticity of native brain parenchyma. The hydrogel is crosslinked by dynamic boronate ester bonds between phenylboronic acid grafted hyaluronic acid (HA-PBA) and dopamine grafted gelatin (Gel-Dopa). This HA-PBA/Gel-Dopa hydrogel could be injected into a lesion cavity in a shear-thinning manner with rapid hemostasis, high tissue adhesion, and efficient self-healing. In an in vivo mouse model of brain lesions, the multi-functional injectable hydrogel is found to support neural cell infiltration, decrease astrogliosis and glial scars, and close the lesions. The results suggest a role for extracellular matrix-mimicking viscoelasticity in brain lesion healing, and motivate additional experimentation in larger animals as the technology progresses toward potential application in humans.

Rational Design of Soft-Hard Interfaces through Bioinspired Engineering.

Soft-hard tissue interfaces in nature present a diversity of hierarchical transitions in composition and structure to address the challenge of stress concentrations that would otherwise arise at their interface. The translation of these into engineered materials holds promise for improved function of biomedical interfaces. Here, soft-hard tissue interfaces found in the body in health and disease, and the application of the diverse, functionally graded, and hierarchical structures that they present to bioinspired engineering materials are reviewed. A range of such bioinspired engineering materials and associated manufacturing technologies that are on the horizon in interfacial tissue engineering, hydrogel bioadhesion at the interfaces, and healthcare and medical devices are described.

A Fenestrated Balloon Expandable Stent System for the Treatment of Aortoiliac Occlusive Disease.

In aortoiliac occlusive disease, atherosclerotic plaques can occlude the distal aortic bifurcation and proximal bilateral iliac artery and thus cause ischemia in the lower extremity. This is typically treated by restoring patency with balloon expandable stents. Stents are typically deployed in a "kissing stent" configuration into the bilateral iliac arteries and into the distal aortic bifurcation lumen to restore antegrade arterial flow. However, these stents typically become re-occluded by plaques. To understand the reasons for this and look for solutions, we simulated flow dynamics in the aortic bifurcation in the presence and absence of stents using computational fluid dynamics. Results demonstrated that the kissing stent configuration was associated with high levels of vorticity and flow constriction. These prothrombotic variables were alleviated in an alternative, aortoiliac fenestrated (AIFEN), tapered, and balloon-expandable stent design. Our findings suggest that stent design can be tailored to improve flow fields for aortoiliac stenting.

The fibrous character of pericellular matrix mediates cell mechanotransduction.

Cells in solid tissues sense and respond to mechanical signals that are transmitted through extracellular matrix (ECM) over distances that are many times their size. This long-range force transmission is known to arise from strain-stiffening and buckling in the collagen fiber ECM network, but must also pass through the denser pericellular matrix (PCM) that cells form by secreting and compacting nearby collagen. However, the role of the PCM in the transmission of mechanical signals is still unclear. We therefore studied an idealized computational model of cells embedded within fibrous collagen ECM and PCM. Our results suggest that the smaller network pore sizes associated with PCM attenuates tension-driven collagen-fiber alignment, undermining long-range force transmission and shielding cells from mechanical stress. However, elongation of the cell body or anisotropic cell contraction can compensate for these effects to enable long distance force transmission. Results are consistent with recent experiments that highlight an effect of PCM on shielding cells from high stresses. Results have implications for the transmission of mechanical signaling in development, wound healing, and fibrosis.

Quantification of the flexural rigidity of endovascular surgical devices using three-point bending tests.

Endovascular surgical procedures require the navigation of catheters and wires through the vasculature to reach distal target sites. Quantitative frameworks for device selection hold the potential to improve the tracking of endovascular devices through vascular anatomy by personalizing the device flexural rigidity to an individual's anatomy. However, data are lacking to facilitate this technology, in part because typical endovascular devices have intricate spatial variations in mechanical properties that are challenging and tedious to quantify repeatably. We therefore developed a three-point bend test methodology using a custom rig and applied it to measure lengthwise flexural rigidity profiles of endovascular devices that are used to target the cerebral vasculature. The methodology demonstrated high repeatability and was able to characterize transition zones. We applied the methodology to generate the first comprehensive, quantitative library of device flexural rigidities, spanning guidewires, intermediate guides, and long sheaths. We observed that these three classes of device have properties that fall into distinct ranges. Additional plots examining relationships between flexural rigidity, device diameter, and length revealed application-specific trends in flexural properties. This methodology and the data allow for standardized characterization and comparisons to aid device selection, and have the potential to both enhance surgical planning and inform future innovation.