Journal of Otolaryngology and Ophthalmology of Shandong University ›› 2026, Vol. 40 ›› Issue (3): 7-15.doi: 10.6040/j.issn.1673-3770.0.2025.177

• Original Article • Previous Articles     Next Articles

Packaging of AAV-ie-Tnfaip8l2 and its effect on improving auditory function in Tnfaip8l2-/- mice

WANG Min, LIU Hao, JIANG Luhan, TAN Liyue, LI Wen, FU Xiaolong   

  1. Medical Science and Technology Innovation Center, Shandong First Medical University & Shandong Academy of Medical Sciences, Jinan 250117, Shandong, China
  • Published:2026-05-22

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Abstract: Objective Adeno-associated virus(AAV)serves as a critical vector for gene therapy. This study aimed to package AAV-ie-Tnfaip8l2 virus, systematically evaluate whether AAV could be used as a vector to achieve in situ overexpression of Tnfaip8l2 protein in the cochlea, and explore its transduction efficiency in the inner ear tissues of gene knockout mouse models and therapeutic effects on auditory function. Methods The recombinant expression vector AAV-ie-Tnfaip8l2 was constructed using molecular cloning techniques. AAV-ie capsid plasmids and helper plasmids were co-transfected into 293T cells to complete the assembly and replication of viral particles. High-purity, high-titer recombinant AAV preparations were obtained via ultracentrifugation and chromatography purification. Using microinjection technology, AAV-ie-Tnfaip8l2 was injected into the round window of newborn Tnfaip8l2-/- mice through the round window membrane. When the Tnfaip8l2-/-mice reached 35 days of age(the typical period when hearing loss becomes apparent), the therapeutic effects were evaluated using methods such as audiological tests, scanning electron microscopy, and immunofluorescence. Results High-titer AAV-ie-Tnfaip8l2 virus could be successfully packaged for cochlear round window injection, enabling efficient in situ overexpression of the Tnfaip8l2 in the cochlea of Tnfaip812-/- mice. Audiological function tests showed that compared with the control groups, the click, tone burst(4 k, 8 k, 12 k, 16 k, 24 k, 32 k), and DPOAE thresholds in the AAV-ie-Tnfaip8l2 replenishment group mice were significantly reduced(P<0.05, n=3). Scanning electron microscopy revealed that the disordered arrangement of stereociliary bundles in outer hair cells was significantly improved. Conclusion This study confirmed that AAV-ie-Tnfaip8l2 can achieve efficient in situ expression of the Tnfaip8l2 in the mouse cochlea via round window injection, partially rescuing the hearing function of Tnfaip8l2-/- mice. These findings provide new experimental evidence for therapeutic strategies for age-related hearing loss.

Key words: Adeno-associated virus, Hearing loss, TNFAIP8L2, Gene function compensation

CLC Number: 

  • R764
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