A microdevice platform for characterizing the effect of mechanical strain magnitudes on the maturation of iPSC-Cardiomyocytes.

The use of human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) as an in vitro model of the heart is limited by their structurally and functionally immature phenotypes. During heart development, mechanical stimuli from in vivo microenvironments are known to regulate cardiomyocyte gene expression and maturation. Accordingly, protocols for culturing iPSC-CMs have recently incorporated mechanical or electromechanical stimulation to induce cellular maturation in vitro; however, the response of iPSC-CMs to different mechanical strain magnitudes is unknown, and existing techniques lack the capability to dynamically measure changes to iPSC-CM contractility in situ as maturation progresses. We developed a microdevice platform which applies cyclical strains of varying magnitudes (5%, 10%, 15% and 20%) to a monolayer of iPSC-CMs, coincidentally measuring contractile stress during mechanical stimulation using fluorescent nanobeads embedded in the microdevice's suspended membrane. Cyclic strain was found to induce circumferential cell alignment on the actuated membranes. In situ contractility measurements revealed that cyclic stimulation gradually increased cardiomyocyte contractility during a 10-day culture period. The contractile stress of iPSC-CM monolayers was found to increase with a higher strain magnitude and plateaued at 15% strain. Cardiomyocyte contractility positively correlated with the elongation of sarcomeres and an increased expression of ß-myosin heavy chain (MYH7) in a strain magnitude-dependent manner, illustrating how mechanical stress can be optimized for the phenotypic and proteomic maturation of the cells. iPSC-CMs with improved maturity have the potential to create a more accurate heart model in vitro for applications in disease modeling and therapeutic discovery.

Stiffness and ATP recovery of stored red blood cells in serum.

In transfusion medicine, there has been a decades-long debate about whether the age of stored red blood cells (RBCs) is a factor in transfusion efficacy. Existing clinical studies investigating whether older RBCs cause worse clinical outcomes have provided conflicting information: some have shown that older blood is less effective, while others have shown no such difference. The controversial results could have been biased by the vastly different conditions of the patients involved in the clinical studies; however, another source of inconsistency is a lack of understanding of how well and quickly stored RBCs can recover their key parameters, such as stiffness and ATP concentration, after transfusion. In this work, we quantitatively studied the stiffness and ATP recovery of stored RBCs in 37 °C human serum. The results showed that in 37 °C human serum, stored RBCs are able to recover their stiffness and ATP concentration to varying extents depending on how long they have been stored. Fresher RBCs (1-3 weeks old) were found to have a significantly higher capacity for stiffness and ATP recovery in human serum than older RBCs (4-6 weeks old). For instance, for 1-week-old RBCs, although the shear modulus before recovery was 1.6 times that of fresh RBCs, 97% of the cells recovered in human serum to have 1.1 times the shear modulus of fresh RBCs, and the ATP concentration of 1-week-old RBCs after recovery showed no difference from that of fresh RBCs. However, for 6-week-old RBCs, only ~70% of the RBCs showed stiffness recovery in human serum; their shear modulus after recovery was still 2.1 times that of fresh RBCs; and their ATP concentration after recovery was 25% lower than that of fresh RBCs. Our experiments also revealed that the processes of stiffness recovery and ATP recovery took place on the scale of tens of minutes. We hope that this study will trigger the next steps of comprehensively characterizing the recovery behaviors of stored RBCs (e.g., recovery of normal 2,3-DPG [2,3-Diphosphoglycerate]and SNO [S-nitrosation] levels) and quantifying the in vivo recovery of stored RBCs in transfusion medicine.

Microdevice Platform for Continuous Measurement of Contractility, Beating Rate, and Beating Rhythm of Human-Induced Pluripotent Stem Cell-Cardiomyocytes inside a Controlled Incubator Environment.

The heart completes a complex set of tasks, including the initiation or propagation of an electrical signal with regularity (proper heart rate and rhythm) and generating sufficient force of contraction (contractility). Probing mechanisms of heart diseases and quantifying drug efficacies demand a platform that is capable of continuous operation inside a cell incubator for long-term measurement of cardiomyocyte (CM) monolayers. Here, we report a microdevice array that is capable of performing continuous, long-term (14 days) measurement of contractility, beating rate, and beating rhythm in a monolayer of human-induced pluripotent stem cell-CMs (hiPSC-CMs). The device consists of a deformable membrane with embedded carbon nanotube (CNT)-based strain sensors. Contraction of the hiPSC-CMs seeded on the membrane induces electrical resistance change of the CNT strain sensor. Continuously reading the sensor signals revealed that hiPSC-CMs started to beat from day 2 and plateaued on day 5. Average contractile stress generated by a monolayer of hiPSC-CMs was determined to be 2.34 ± 0.041 kPa with a beating rate of 1.17 ± 0.068 Hz. The device arrays were also used to perform comprehensive measurement of the beating rate, rhythm, and contractility of the hiPSC-CMs and quantify the cell responses to different concentrations of agonists and antagonists, which altered the average contractile stress to the range of 1.15 ± 0.13 to 3.96 ± 0.53 kPa. The continuous measurement capability of the device arrays also enabled the generation of Poincaré plots for revealing subtle changes in the beating rhythm of hiPSC-CMs under different drug treatments.

Mechanical stability of the cell nucleus - roles played by the cytoskeleton in nuclear deformation and strain recovery.

Extracellular forces transmitted through the cytoskeleton can deform the cell nucleus. Large nuclear deformations increase the risk of disrupting the integrity of the nuclear envelope and causing DNA damage. The mechanical stability of the nucleus defines its capability to maintain nuclear shape by minimizing nuclear deformation and allowing strain to be minimized when deformed. Understanding the deformation and recovery behavior of the nucleus requires characterization of nuclear viscoelastic properties. Here, we quantified the decoupled viscoelastic parameters of the cell membrane, cytoskeleton, and the nucleus. The results indicate that the cytoskeleton enhances nuclear mechanical stability by lowering the effective deformability of the nucleus while maintaining nuclear sensitivity to mechanical stimuli. Additionally, the cytoskeleton decreases the strain energy release rate of the nucleus and might thus prevent shape change-induced structural damage to chromatin.

Moldable elastomeric polyester-carbon nanotube scaffolds for cardiac tissue engineering.

Polymer biomaterials are used to construct scaffolds in tissue engineering applications to assist in mechanical support, organization, and maturation of tissues. Given the flexibility, electrical conductance, and contractility of native cardiac tissues, it is desirable that polymeric scaffolds for cardiac tissue regeneration exhibit elasticity and high electrical conductivity. Herein, we developed a facile approach to introduce carbon nanotubes (CNTs) into poly(octamethylene maleate (anhydride) 1,2,4-butanetricarboxylate) (124 polymer), and developed an elastomeric scaffold for cardiac tissue engineering that provides electrical conductivity and structural integrity to 124 polymer. 124 polymer-CNT materials were developed by first dispersing CNTs in poly(ethylene glycol) dimethyl ether porogen and mixing with 124 prepolymer for molding into shapes and crosslinking under ultraviolet light. 124 polymers with 0.5% and 0.1% CNT content (wt) exhibited improved conductivity against pristine 124 polymer. With increasing the CNT content, surface moduli of hybrid polymers were increased, while their bulk moduli were decreased. Furthermore, increased swelling of hybrid 124 polymer-CNT materials was observed, suggesting their improved structural support in an aqueous environment. Finally, functional characterization of engineered cardiac tissues using the 124 polymer-CNT scaffolds demonstrated improved excitation threshold in materials with 0.5% CNT content (3.6±0.8V/cm) compared to materials with 0% (5.1±0.8V/cm) and 0.1% (5.0±0.7V/cm), suggesting greater tissue maturity. 124 polymer-CNT materials build on the advantages of 124 polymer elastomer to give a versatile biomaterial for cardiac tissue engineering applications. STATEMENT OF SIGNIFICANCE: Achieving a high elasticity and a high conductivity in a single cardiac tissue engineering material remains a challenge. We report the use of CNTs in making electrically conductive and mechanically strong polymeric scaffolds in cardiac tissue regeneration. CNTs were incorporated in elastomeric polymers in a facile and reproducible approach. Polymer-CNT materials were able to construct complicated scaffold structures by injecting the prepolymer into a mold and crosslinking the prepolymer under ultraviolet light. CNTs enhanced electrical conductivity and structural support of elastomeric polymers. Hybrid polymeric scaffolds containing 0.5wt% CNTs increased the maturation of cardiac tissues fabricated on them compared to pure polymeric scaffolds. The cardiac tissues on hybrid polymer-CNT scaffolds showed earlier beating than those on pure polymer scaffolds. In the future, fabricated polymer-CNT scaffolds could also be used to fabricate other electro-active tissues, such neural and skeletal muscle tissues. In the future, fabricated polymer-CNT scaffolds could also be used to fabricate other electro-active tissues, such as neural and skeletal muscle tissues.

Microfluidic Assessment of Frying Oil Degradation.

Monitoring the quality of frying oil is important for the health of consumers. This paper reports a microfluidic technique for rapidly quantifying the degradation of frying oil. The microfluidic device generates monodispersed water-in-oil droplets and exploits viscosity and interfacial tension changes of frying oil samples over their frying/degradation process. The measured parameters were correlated to the total polar material percentage that is widely used in the food industry. The results reveal that the steady-state length of droplets can be used for unambiguously assessing frying oil quality degradation.

Stiffening of sickle cell trait red blood cells under simulated strenuous exercise conditions.

The higher risk of vaso-occlusion events and sudden death for sickle-cell trait (SCT) athletes has been speculatively ascribed to SCT red blood cell (RBC) stiffening during strenuous exercise. However, the microenvironmental changes that could induce the stiffening of SCT RBCs are unknown. To address this question, we measured the mechanical properties of and changes in SCT RBCs under deoxygenated and acidic environments, which are two typical conditions present in the circulation of athletes undertaking strenuous exercise. The results reveal that SCT RBCs are inherently stiffer than RBCs from non-SCT healthy subjects, and a lower pH further stiffens the SCT cells. Furthermore, at both normal and low pH levels, deoxygenation was found to not be the cause of the stiffness of SCT RBCs. This study confirms that the stiffening of SCT RBCs occurs at a low pH and implies that SCT RBC stiffening could be responsible for vaso-occlusion in SCT athletes during strenuous exercise.

Mechanical differences of sickle cell trait (SCT) and normal red blood cells.

Sickle cell trait (SCT) is a condition in which an individual inherits one sickle hemoglobin gene (HbS) and one normal beta hemoglobin gene (HbA). It has been hypothesized that under extreme physical stress, the compromised mechanical properties of the red blood cells (RBCs) may be the underlying mechanism of clinical complications of sickle cell trait individuals. However, whether sickle cell trait (SCT) should be treated as physiologically normal remains controversial. In this work, the mechanical properties (i.e., shear modulus and viscosity) of individual RBCs were quantified using a microsystem capable of precisely controlling the oxygen level of RBCs' microenvironment. Individual RBCs were deformed under shear stress. After the release of shear stress, the dynamic cell recovery process was captured and analyzed to extract the mechanical properties of single RBCs. The results demonstrate that RBCs from sickle cell trait individuals are inherently stiffer and more viscous than normal RBCs from healthy donors, but oxygen level variations do not alter their mechanical properties or morphology.

Traditional chinese medicine prescriptions enhance growth performance of heat stressed beef cattle by relieving heat stress responses and increasing apparent nutrient digestibility.

The present aim was to investigate the effects of traditional Chinese medicine prescriptions (TCM) on body temperature, blood physiological parameters, nutrient apparent digestibility and growth performance of beef cattle under heat stress conditions. Twenty-seven beef cattle were randomly divided into three groups as following; i) high temperature control (HTC), ii) traditional Chinese medicine prescriptions I+high temperature (TCM I) and iii) traditional Chinese medicine prescriptions II+high temperature (TCM II) (n = 9 per group). The results showed that the mean body temperature declined in TCM II treatment (p<0.05). Serum T3 and T4 levels with TCM I and TCM II treatments elevated (p<0.05), and serum cortisol levels of TCM I treatments decreased (p<0.05), compared with the HTC group. Total protein, albumin, globulin in TCM II treatments elevated and blood urea nitrogen levels of both TCM treatments increased, but glucose levels of both TCM treatments decreased, compared with the HTC group (p<0.05). The apparent digestibility of organic matter and crude protein with TCM I treatment increased, and the apparent digestibility of acid detergent fiber elevated in both TCM treatments (p<0.05). Average daily feed intake was not different among three groups, however average daily gain increased and the feed:gain ratio decreased with both TCM treatments, compared with the HTC group (p<0.05). The present results suggest that dietary supplementation with TCM I or TCM II improves growth performance of heat stressed beef cattle by relieving heat stress responses and increasing nutrient apparent digestibility.