Autoantibodies immuno-mechanically modulate platelet contractile force and bleeding risk.

Altered mechanotransduction has been proposed as a putative mechanism for disease pathophysiology, yet evidence remains scarce. Here we introduce a concept we call single cell immuno-mechanical modulation, which links immunology, integrin biology, cellular mechanics, and disease pathophysiology and symptomology. Using a micropatterned hydrogel-laden coverslip compatible with standard fluorescence microscopy, we conduct a clinical mechanobiology study, specifically focusing on immune thrombocytopenia (ITP), an autoantibody-mediated platelet disorder that currently lacks a reliable biomarker for bleeding risk.

Less-deformable erythrocyte subpopulations biomechanically induce endothelial inflammation in sickle cell disease.

Sickle cell disease (SCD) is canonically characterized by reduced red blood cell (RBC) deformability, leading to microvascular obstruction and inflammation. Although the biophysical properties of sickle RBCs are known to influence SCD vasculopathy, the contribution of poor RBC deformability to endothelial dysfunction has yet to be fully explored. Leveraging interrelated in vitro and in silico approaches, we introduce a new paradigm of SCD vasculopathy in which poorly deformable sickle RBCs directly cause endothelial dysfunction via mechanotransduction, during which endothelial cells sense and pathophysiologically respond to aberrant physical forces independently of microvascular obstruction, adhesion, or hemolysis.

Single-pericyte nanomechanics measured by contraction cytometry.

Pericytes line the microvasculature throughout the body and play a key role in regulating blood flow by constricting and dilating vessels. However, the biophysical mechanisms through which pericytes transduce microenvironmental chemical and mechanical cues to mediate vessel diameter, thereby impacting oxygen and nutrient delivery, remain largely unknown. This knowledge gap is clinically relevant as numerous diseases are associated with the aberrant contraction of pericytes, which are unusually susceptible to injury.

An economical in-class sticker microfluidic activity develops student expertise in microscale physics and device manufacturing.

Educating new students in miniaturization science remains challenging due to the non-intuitive behavior of microscale objects and specialized layer-by-layer assembly approaches. In our analysis of the existing literature, we noted that it remains difficult to have low cost activities that elicit deep learning. Furthermore, few activities have stated learning goals and measurements of effectiveness.

Opportunities and considerations for studying liver disease with microphysiological systems on a chip.

Advances in microsystem engineering have enabled the development of highly controlled models of the liver that better recapitulate the unique biological conditions. In just a few short years, substantial progress has been made in creating complex mono- and multi-cellular models that mimic key metabolic, structural, and oxygen gradients crucial for liver function. Here we review: 1) the state-of-the-art in liver-centric microphysiological systems and 2) the array of liver diseases and pressing biological and therapeutic challenges which could be investigated with these systems.

Simple and Economical Extraction of Viral RNA and Storage at Ambient Temperature.

RNA extraction is essential for the molecular detection of common viral pathogens. However, available extraction methods and the need for ultra-cold storage limit molecular testing in resource-constrained settings. Herein, we describe the development of an economical xtraction and torage (RNAES) protocol that eliminates requirements for instrumentation, expensive materials, and preserved cold chain.

Resolving the missing link between single platelet force and clot contractile force.

Blood clot contraction plays an important role in wound healing and hemostasis. Although clot contraction is known to be driven by platelets, how single platelet forces relate to the forces generated by macroscopic clots remains largely unknown. Using our microfabricated high-throughput platelet contraction cytometer, we find that single platelets have an average force of 34 nN ( healthy individuals).

Pathologically stiff erythrocytes impede contraction of blood clots.

Background: Blood clot contraction, volume shrinkage of the clot, is driven by platelet contraction and accompanied by compaction of the erythrocytes and their gradual shape change from biconcave to polyhedral, with the resulting cells named polyhedrocytes.

Objectives: Here, we examined the role of erythrocyte rigidity on clot contraction and erythrocyte shape transformation.

Methods: We used an optical tracking methodology that allowed us to quantify changes in contracting clot size over time.

Platelet heterogeneity enhances blood clot volumetric contraction: An example of asynchrono-mechanical amplification.

Physiological processes such as blood clotting and wound healing as well as pathologies such as fibroses and musculoskeletal contractures, all involve biological materials composed of a contracting cellular population within a fibrous matrix, yet how the microscale interactions among the cells and the matrix lead to the resultant emergent behavior at the macroscale tissue level remains poorly understood. Platelets, the anucleate cell fragments that do not divide nor synthesize extracellular matrix, represent an ideal model to study such systems. During blood clot contraction, microscopic platelets actively pull fibers to shrink the macroscale clot to less than 10% of its initial volume.

Vascularized Microfluidics and Their Untapped Potential for Discovery in Diseases of the Microvasculature.

Microengineering advances have enabled the development of perfusable, endothelialized models of the microvasculature that recapitulate the unique biological and biophysical conditions of the microcirculation in vivo. Indeed, at that size scale (<100 μm)-where blood no longer behaves as a simple continuum fluid; blood cells approximate the size of the vessels themselves; and complex interactions among blood cells, plasma molecules, and the endothelium constantly ensue-vascularized microfluidics are ideal tools to investigate these microvascular phenomena. Moreover, perfusable, endothelialized microfluidics offer unique opportunities for investigating microvascular diseases by enabling systematic dissection of both the blood and vascular components of the pathophysiology at hand.

Stiffness based enrichment of leukemia cells using microfluidics.

To improve the survival rate of cancer patients, new diagnosis strategies are necessary to detect lower levels of cancer cells before and after treatment regimens. The scarcity of diseased cells, particularly in residual disease after treatment, demands highly sensitive detection approaches or the ability to enrich the diseased cells in relation to normal cells. We report a label-free microfluidic approach to enrich leukemia cells from healthy cells using inherent differences in cell biophysical properties.

Label-free hematology analysis using deep-ultraviolet microscopy.

Hematological analysis, via a complete blood count (CBC) and microscopy, is critical for screening, diagnosing, and monitoring blood conditions and diseases but requires complex equipment, multiple chemical reagents, laborious system calibration and procedures, and highly trained personnel for operation. Here we introduce a hematological assay based on label-free molecular imaging with deep-ultraviolet microscopy that can provide fast quantitative information of key hematological parameters to facilitate and improve hematological analysis. We demonstrate that this label-free approach yields 1) a quantitative five-part white blood cell differential, 2) quantitative red blood cell and hemoglobin characterization, 3) clear identification of platelets, and 4) detailed subcellular morphology.

The biophysics and mechanics of blood from a materials perspective.

Cells actively interact with their microenvironment, constantly sensing and modulating biochemical and biophysical signals. Blood comprises a variety of non-adherent cells that interact with each other and with endothelial and vascular smooth muscle cells of the blood vessel walls. Blood cells are further experiencing a range of external forces by the hemodynamic environment and they also exert forces to remodel their local environment.

Getting a good view: imaging of platelets under flow.

As the anucleate cells responsible for hemostasis and thrombosis, platelets are exposed to a myriad of biophysical and biochemical stimuli within vasculature and heterogeneous blood clots. Highly controlled, reductionist imaging studies have been instrumental in providing a detailed and quantitative understanding of platelet biology and behavior, and have helped elucidate some surprising functions of platelets. In this review, we highlight the tools and approaches that enable visualization of platelets in conjunction with precise control over the local biofluidic and biochemical microenvironment.

Feeling the Force: Measurements of Platelet Contraction and Their Diagnostic Implications.

In addition to the classical biological and biochemical framework, blood clots can also be considered as active biomaterials composed of dynamically contracting platelets, nascent polymeric fibrin that functions as a matrix scaffold, and entrapped blood cells. As platelets sense, rearrange, and apply forces to the surrounding microenvironment, they dramatically change the material properties of the nascent clot, increasing its stiffness by an order of magnitude. Hence, the mechanical properties of blood clots are intricately tied to the forces applied by individual platelets.

Smartphone app for non-invasive detection of anemia using only patient-sourced photos.

We introduce a paradigm of completely non-invasive, on-demand diagnostics that may replace common blood-based laboratory tests using only a smartphone app and photos. We initially targeted anemia, a blood condition characterized by low blood hemoglobin levels that afflicts >2 billion people. Our app estimates hemoglobin levels by analyzing color and metadata of fingernail bed smartphone photos and detects anemia (hemoglobin levels <12.

A microengineered vascularized bleeding model that integrates the principal components of hemostasis.

Hemostasis encompasses an ensemble of interactions among platelets, coagulation factors, blood cells, endothelium, and hemodynamic forces, but current assays assess only isolated aspects of this complex process. Accordingly, here we develop a comprehensive in vitro mechanical injury bleeding model comprising an "endothelialized" microfluidic system coupled with a microengineered pneumatic valve that induces a vascular "injury". With perfusion of whole blood, hemostatic plug formation is visualized and "in vitro bleeding time" is measured.

Microfluidic Transduction Harnesses Mass Transport Principles to Enhance Gene Transfer Efficiency.

Ex vivo gene therapy using lentiviral vectors (LVs) is a proven approach to treat and potentially cure many hematologic disorders and malignancies but remains stymied by cumbersome, cost-prohibitive, and scale-limited production processes that cannot meet the demands of current clinical protocols for widespread clinical utilization. However, limitations in LV manufacture coupled with inefficient transduction protocols requiring significant excess amounts of vector currently limit widespread implementation. Herein, we describe a microfluidic, mass transport-based approach that overcomes the diffusion limitations of current transduction platforms to enhance LV gene transfer kinetics and efficiency.

Magnetic forces enable controlled drug delivery by disrupting endothelial cell-cell junctions.

The vascular endothelium presents a major transport barrier to drug delivery by only allowing selective extravasation of solutes and small molecules. Therefore, enhancing drug transport across the endothelial barrier has to rely on leaky vessels arising from disease states such as pathological angiogenesis and inflammatory response. Here we show that the permeability of vascular endothelium can be increased using an external magnetic field to temporarily disrupt endothelial adherens junctions through internalized iron oxide nanoparticles, activating the paracellular transport pathway and facilitating the local extravasation of circulating substances.

Platelet-Microcapsule Hybrids Leverage Contractile Force for Targeted Delivery of Hemostatic Agents.

We report a cell-mediated, targeted drug delivery system utilizing polyelectrolyte multilayer capsules that hybridize with the patient's own platelets upon intravenous administration. The hybridized platelets function as the sensor and actuator for targeted drug delivery and controlled release in our system. These capsules are biochemically and mechanically tuned to enable platelet adhesion and capsule rupture upon platelet activation and contraction, enabling the targeted and controlled "burst" release of an encapsulated biotherapeutic.

Single-platelet nanomechanics measured by high-throughput cytometry.

Haemostasis occurs at sites of vascular injury, where flowing blood forms a clot, a dynamic and heterogeneous fibrin-based biomaterial. Paramount in the clot's capability to stem haemorrhage are its changing mechanical properties, the major drivers of which are the contractile forces exerted by platelets against the fibrin scaffold. However, how platelets transduce microenvironmental cues to mediate contraction and alter clot mechanics is unknown.

Cellular softening mediates leukocyte demargination and trafficking, thereby increasing clinical blood counts.

Leukocytes normally marginate toward the vascular wall in large vessels and within the microvasculature. Reversal of this process, leukocyte demargination, leads to substantial increases in the clinical white blood cell and granulocyte count and is a well-documented effect of glucocorticoid and catecholamine hormones, although the underlying mechanisms remain unclear. Here we show that alterations in granulocyte mechanical properties are the driving force behind glucocorticoid- and catecholamine-induced demargination.