Cyclical Stretching Induces Excess Intracellular Ca2+ Influx in Human Keloid-Derived Fibroblasts In Vitro.

BACKGROUND: The incidence of keloids is higher in the case of darker skin. It is more common in the parts exposed to stretching (thorax, abdomen, and joints). Cyclical stretching reportedly induced each Ca2+ spike through differential mechanosensitive channels in human synovial and dermal fibroblasts. Therefore, the authors hypothesized that cyclical stretching also induces a specific Ca2+ spike in keloid-derived fibroblasts. METHODS: This in vitro study compared the intracellular calcium dynamics induced by cyclical stretching between control (human dermal fibroblasts) and keloid (human keloid-derived fibroblasts) groups. Each group was exposed to two-dimensional stretch using an originally developed stretch microdevice. Intracellular Ca2+ was observed for 5 minutes, including 30 seconds of baseline, under a fluorescent confocal laser microscope. The intracellular Ca2+ concentration was evaluated every 0.5 second using the fluorescence intensity ratio. A positive cellular response was defined as a rise of the ratio by greater than or equal to 20%. The normal response cutoff value was determined by receiver operating characteristic analysis. RESULTS: The keloid groups were significantly more responsive than the control groups (15.7% versus 8.2%; P = 0.029). In the cellular response-positive cells, the keloid groups reached significantly higher intracellular Ca2+ concentration peaks than the control groups (2.20 versus 1.26; P = 0.0022). The cutoff value was 1.77, and 10.4% of the keloid-derived fibroblasts exhibited a hyper-Ca2+ spike above the normal range. CONCLUSIONS: Keloid-derived fibroblasts with a hyper-Ca2+ spike might constitute a keloid-specific subpopulation. Hereafter, the authors will study whether the normalization of excessive intracellular Ca2+ concentration leads to keloid treatment in vivo. CLINICAL RELEVANCE STATEMENT: This study result provided a clue to the onset mechanism of keloids, which the authors hope will lead to the development of new therapy in the future.

Control of microtubule trajectory within an electric field by altering surface charge density.

One of challenges for using microtubules (MTs) driven by kinesin motors in microfluidic environments is to control their direction of movement. Although applying physical biases to rectify MTs is prevalent, it has not been established as a design methodology in conjunction with microfluidic devices. In the future, the methodology is expected to achieve functional motor-driven nanosystems. Here, we propose a method to guide kinesin-propelled MTs in multiple directions under an electric field by designing a charged surface of MT minus ends labeled with dsDNA via a streptavidin-biotin interaction. MTs labeled with 20-bp or 50-bp dsDNA molecules showed significantly different trajectories according to the DNA length, which were in good agreement with values predicted from electrophoretic mobilities measured for their minus ends. Since the effective charge of labeled DNA molecules was equal to that of freely dispersed DNA molecules in a buffer solution, MT trajectory could be estimated by selecting labeling molecules with known charges. Moreover, the estimated trajectory enables to define geometrical sizes of a microfluidic device. This rational molecular design and prediction methodology allows MTs to be guided in multiple directions, demonstrating the feasibility of using molecular sorters driven by motor proteins.