Research On Atohl Gene Transduction To Guinea Pig Cochlear Cells Mediated By HA-PEI Nanoparticle Vectors

Since the first experiment aimed at developing a shiverer mouse model for correcting hearing disorders resulting from genetic abnormalities of the central nervous system (CNS) by Fujiyoshi in 1994, researches on gene therapy for sensorineural deafness have been greatly development.Sensorineural hearing loss is largely caused by the degeneration of the cochlea, a sensory organ for hearing. Studies of human temporal bones have revealed that SNHL can be caused by the degeneration of various components of the cochlea. Among the various types of cochlear cells, hair cells, which convert sound stimuli to neural signals, have been a central therapeutic target for developing novel therapeutics. In mammals, the cochlear components described above have virtually no regenerative capacity. However, recent studies have demonstrated that Atonal homologl (Atohl)/Mathl is an essential for hair cell (HC) differentiation and ectopic hair cells were found after/Mathl plasmid were successfully transfected to the cultured Corti. Our research group has reported that after transfecting the Atohl gene by an adenovirus gene vector to noise-induced hearing loss guinea pig cochlea, the damaged/lost stereocilia bundles were repaired or regenerated, which promoted the functional restoration. However, adenovirus gene vector has strong host antiviral response and performed as transient transfection, which makes it not an optimal vector for gene delivery and difficult to be authorize by the clinical ethics committee. Furthermore, being different from the purpose of killing cells in tumor cells’ gene therapy research, the research on ear disease is to promote more cell regeneration or repaired. Therefore, in order to screen a better nonviral vectors for ear disease, based on the previous work of our research group, we synthesized a new nanoparticle gene vector to transfer the Atohl gene plasmid into health and noise induced hearing loss guinea pig cochlea. Then, we studied the transfer efficiency and biological effects of new gene vector using the auditory function measurement and morphological detection.In the first part of this experiment, we used Hyaluronic acid (HA) to modified polyethylenimine(PEI) nanoparticles to obtain a HA-PEI nanoparticle for gene delivery. Then, we put the HA-PEI-Atohl and PEI-Atohl nano gene complexes separately onto the round window membrane through an ear surgery and compared the different transfection efficiency of the two vectors. In the second part, we used the synthesized nano gene vector to transfer Atohl gene plasmid into cultured hair cells isolated from guinea pigs, and observed the methods, time needed and position of HA-PEI-DNA nano gene complex transferring into hair cells. In the third part, based on the noise-induced hearing loss animal model of guinea pig, we transferred the HA-PEI-Atohl nano gene complexes into deaf guinea pig to observe if there is any therapeutic and preventive effect of Atohl gene, which would provide an option of vector for gene therapy for sensorineural hearing loss.