In-vitro human epidermal keratinocyte migration via dielectrophoresis technique for stimulating chronic wound epithelialization

Abstract

Diabetes patients are at risk of having chronic wounds, which would take months to years to resolve naturally. Unhealed injuries increase the economic burden and clinical complications, which may develop a challenge to the medical community. International diabetic federation statistics report that diabetes will affect 537 million people worldwide in 2021 and is expected to be 643 million by 2030 and 783 million by 2045. Conventional wound care techniques experience challenges due to the device complexity, high cost, unfavourable side effects, painful treatment, and the lengthy wound healing process. To improve wound healing, we must pay attention and take immediate action on the better solution to speed up the wound healing process. In this study, chronic wounds can be countered using the exogenous electrical stimulation technique (EST) by dielectrophoresis (DEP). The advantages DEP technique is label-free, highly sensitive, and selective for particle/cell migration trajectory. In human skin, the epidermis is the outermost layer of the skin and serves as a body shield. The human epidermal keratinocyte cells (HEK) are the building block of the skin epidermis and are crucial for the re-epithelization of the skin. This study focuses on the rapid in-vitro migration of HEK based on DEP force (FDEP) for expeditious epithelialization of chronic wounds. We began by validating polystyrene (PS) particles to predict the behaviour of HEK to estimate their high-speed migration rate of the NDEP response for rapid migration. Because the size of the HEK varies, we used proof-of-concept PS particles 9.9 and 14.6 μm in this study. The MyDEP is a standalone software designed to study dielectric dynamic change and cell response to an alternating current (AC) electric field. Finite element method (FEM), COMSOL Multiphysics 5.6, was utilized to compute the electric field intensity distribution and particle trajectory based on DEP and drag forces in 2-D and 3-D. The DEP microelectrode was designed with AutoCAD and fabricated via surface micromachining technique to produce 40 and 60 μm ratchet and parallel microelectrodes. The DEP experiment was performed by applying sinusoidal wave alternating current (AC) potential at the peak-to-peak voltages of 8, 10, and 12 VPP in a fabricated microelectrode from 100 kHz to 25 MHz. In experiments, the dominant NDEP drives HEK cells to the local electric field minima in the region of interest. The motion analysis software (DIPP-MotionV) was used to track cell migration and estimates the particles and cell's speed. The DEP experimental result's migration velocity and DEP response are validated and correlated with the FEM and MyDEP data. In the traditional in-vitro migration assays, such as scratch and microfluidics-based system assays, the average velocity to close the wound is 0.007 to 0.003 μm/s, respectively. Compared to the conventional technique, the current study successfully migrates in vitro HEK cells at a rate of 263 μm/s in 5 MHz, 8 VPP in the 40 μm ratchet microelectrode. Additionally, we compared the device performance among 40 and 60 μm ratchet and parallel microelectrode for highly effective particle and cell migration. As a result, the ratchet microelectrode achieved the HEK cell migration efficiency of ~83.67 to 95.05 %. Furthermore, HEK cell viability is examined after applying an electric field, and the cells remain viable within 60 s of exposure. This current study has a novel FEM computation similar to the experiment, research methodology, and core idea of utilizing the DEP method to enhance HEK cell migration for rapid wound epithelization. Therefore, the EST-DEP technique for in-vitro HEK cell migration by 40 μm ratchet microelectrode was efficient, making it suitable for wound epithelization applications for rapid chronic wound closure.