As the development of physical plasma technology, cold atmospheric plasmas (CAPs) have been widely investigated in decontamination, cancer treatment, wound healing, root canal treatment, etc., forming a new research field named plasma medicine. Being a mixture of electrical, chemical, and biological reactive species, CAPs have shown their abilities to enhance nerve stem cells differentiation both in vitro and in vivo and are becoming a promising way for neurological disease treatment in the future. The much more exciting news is that using CAPs may realize one-step, and safely directed, differentiation of neural stem cells (NSCs) for tissue transportation. We demonstrate here the detailed experimental protocol of using a self-made CAP jet device to enhance NSC differentiation in C17.2-NSCs and primary rat neural stem cells, as well as observing the cell fate by inverted and fluorescence microscopy. With the help of immunofluorescence staining technology, we found both the NSCs showed an accelerated differential rate than the untreated group, and ~75% of the NSCs selectively differentiated into neurons, which are mainly mature, cholinergic, and motor neurons.

In the past 20 years, cold atmospheric pressure plasmas (CAPs) have become a new promising way for many biomedical applications, such as disinfection, cancer treatment, root canal treatment, wound healing, and other medical applications. Among these applications, investigations of plasma for skin wound healing has gained huge success both in vitro and in vivo experiments, and also the mechanism behind it has been studied by many groups. In this chapter, we summarize the state-of-the-art progress in wound healing by CAPs. The plasma devices developed for wound healing, the interactions between plasmas and microorganisms/cells/tissues, the in vitro and in vivo treatments, the clinical trials, and biosafety issues are all included.

Cold atmospheric plasmas (CAPs) have been shown to be capable of enhancing stem cell differentiation, especially directed differentiation of neural stem cells (NSCs). Consequently, one-step CAP treatment shows promise as an aid to tissue transplantation. However, the mechanisms involved in the enhancement of NSCs differentiation by CAP treatment are not yet fully understood. We have previously shown that in atmospheric helium plasma jet treatment, nitric oxide (NO) is the main factor involved in promoting NSC differentiation. This article further investigated the possible signaling pathways stimulated by NO in the neuronal differentiation of C17.2-NSCs after plasma treatment. Extracellular and intracellular NO concentrations were measured at different time points of incubation to monitor NO production. Meanwhile, the expressions of related genes and proteins were detected by quantitative real-time polymerase chain reaction and western blot, respectively. It is found that plasma treatment could both generate extracellular NO and increase extracellular NO concentration by inducing inducible nitric oxide synthase expression. The synergetic effect of extracellular and intracellular NO then downregulated Notch1 and Id2, and upregulated Ngn2 and Ascl1, thereby activating downstream NeuroD expression and finally enhancing and directing differentiation of NSCs into neurons.