Scanning Ion Conductance Microscopy Study Reveals the Disruption of the Integrity of the Human Cell Membrane Structure by Oxidative DNA Damage.

Oxidative stress can damage organs, tissues, and cells through reactive oxygen species (ROS) by oxidizing DNA, proteins, and lipids, thereby resulting in diseases. However, the underlying molecular mechanisms remain to be elucidated. In this study, employing scanning ion conductance microscopy (SICM), we explored the early responses of human embryonic kidney (HEK293H) cells to oxidative DNA damage induced by potassium chromate (K2CrO4). We found that the short term (1-2 h) exposure to a low concentration (10 µM) of K2CrO4 damaged the lipid membrane of HEK293H cells, resulting in structural defects and depolarization of the cell membrane and reducing cellular secretion activity shortly after the treatment. We further demonstrated that the K2CrO4 treatment decreased the expression of the cytoskeleton protein, ß-actin, by inducing oxidative DNA damage in the exon 4 of the ß-actin gene. These results suggest that K2CrO4 caused oxidative DNA damage in cytoskeleton genes such as ß-actin and reduced their expression, thereby disrupting the organization of the cytoskeleton beneath the cell membrane and inducing cell membrane damages. Our study provides direct evidence that oxidative DNA damage disrupted human cell membrane integrity by deregulating cytoskeleton gene expression.

Extrusion 3D Printing of Porous Silicone Architectures for Engineering Human Cardiomyocyte-Infused Patches Mimicking Adult Heart Stiffness.

Cardiac patches, three-dimensional (3D) constructs of polymer scaffold and heart muscle cells, have received widespread attention for regenerative therapy to repair damaged heart tissue. The implanted patches should mimic the micromechanical environment of native myocardium for effective integration and optimum mechanical function. In this study, we engineered compliant silicone scaffolds infused with cardiomyocytes (CMs) differentiated from human-induced pluripotent stem cells. Porous scaffolds are fabricated by extrusion 3D printing of room-temperature-vulcanized (RTV) silicone rubber. The stiffness and strength of scaffolds are tailored by designing a polymer strand arrangement during 3D printing. Single-strand scaffold design is found to display a tensile Young's modulus of ∼280 kPa, which is optimum for supporting CMs without impairing their contractility. Uniform distribution of cells in the scaffold is observed, ascribed to 3D migration facilitated by interconnected porous architecture. The patches demonstrated synchronized contraction 10 days after seeding scaffolds with CMs. Indentation measurements reveal that the contracting cell-scaffold patches display local moduli varying from ∼270 to 530 kPa, which covers the upper spectrum of the stiffness range displayed by the human heart. This study demonstrates the effectiveness of a porous 3D scaffold composed of flexible silicone rubber for CMs percolation, supporting a contractile activity, and mimicking native heart stiffness.