The Mechanical Agitation Method of Gene Transfer for Ex-Vivo Gene Therapy

Gene therapy is a therapeutic method used to treat diseases by altering genes within a patient’s cells. The concept of gene therapy emerged as molecular biology evolved from the mere discipline of studying DNA molecules to the scientific art of virtually manipulating the genes of cells. Explosive worldwide research was conducted after the first introduction of the concept of gene therapy into the scientific community. The original aim was to directly modify patient genes through in vivo gene therapeutic approaches. However, DNA molecules introduced into the body by in vivo gene therapy are delivered at a very low frequency into terminally differentiated tissue cells, which typically do not have the capability of self-renewal (Tenenbaum et al., 2003). Because of the short-lived nature of in vivo gene therapy, a defective gene in patients is only temporarily corrected by in vivo gene therapy (Kaloss et al., 1999). The development of gene therapeutic methods in which the corrected patient gene remains permanently has been actively pursued. Ex vivo gene therapeutic methods have been considered as alternative options to gene therapy to overcome the short-lived nature of the corrected genes of in vivo gene therapy. In ex vivo gene therapy, the surgically removed adult stem cells, such as mesenchymal stem cells or hematopoietic stem cells, are typically cultured in a laboratory apparatus. The therapeutic DNA molecules are introduced into the isolated cells, and these transfected cells are then introduced into the patients. By using adult stem cells in ex vivo gene therapeutic methods, the corrected genes that are introduced are, in most cases, expressed permanently once they are corrected properly because the adult stem cells have the capability of selfrenewal (Dube & Denis, 1995; Muller-Sieburg & Sieburg, 2006; Tseng et al., 2006; Nehlin & Barington 2009). In ex vivo gene therapy, genetic manipulation is conducted in a lab outside of the body. However, normal somatic cells, including adult stem cells, do not propagate indefinitely and are vulnerable to epigenetic modification. Therefore, long-term cultures of somatic cells isolated from the body are very difficult to sustain (Beyer & Da sliva, 2006; Tonti & Mannello, 2008). This means that the long-term culture of adult stem cells in ex vivo gene therapy should be avoided as much as possible. Therefore, it is absolutely necessary to deliver therapeutic DNA molecules into isolated cells immediately with high efficiency.