Development of a method for in vitro cultivation of Corti's organ explants using polyethyleneimine

The aim. To develop a method for in vitro cultivation of Corti’s organ explants using polyethyleneimine (PEI) as an adhesive substrate. The study evaluated the potential of PEI as an effective alternative to existing substrates for in vitro cultivation of inner ear samples.

Materials and methods. Explants of Corti’s organ from newborn mice (P1–P2) were used. Glass culture surfaces were treated with a PEI solution followed by treatment with fetal bovine serum. To assess the suitability of the developed explant model for ototoxicity studies, treatment with gentamicin was performed. Isoliquiritigenin was used as an otoprotector. Morphological analysis of images obtained by confocal fluorescence microscopy was conducted. The survival of hair cells after staining with phalloidin labeled with Alexa Fluor 488 was evaluated. The magnitude of stereociliary angles of outer hair cells was assessed.

Results. PEI- coating provided reliable fixation of Corti’s organ explants (up to 72 h) without signs of detachment from the culture surface. The samples maintained their attachment strength through two media changes and subsequent 16 washing cycles during fluorescent staining. Morphological analysis of the control group revealed no signs of toxic effects of PEI on hair cells. The organotypic model using PEI coating reproduced the classic pattern of gentamicin-induced ototoxicity with a basal-apical gradient of damage. For the first time, this model demonstrated the otoprotective effect of isoliquiritigenin (1 μM) against gentamicin-induced ototoxicity (500 μM), most pronounced in the middle turn of the cochlea for both inner and outer hair cells (p < 0.0001 for both comparisons with the gentamicin group).

Conclusion. The efficacy of PEI as an alternative substrate for cultivating inner ear samples has been demonstrated for the first time, showing optimal adhesive properties and no inherent toxicity. The developed method represents a promising approach for ototoxicity research and otoprotector screening.

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