本年度の実験では，内耳に遺伝子導入することを最終目標として下記の2つの実験を実行した．
(1) 超音波照射によるAAVの徐放
AAVをハイドロゲルマイクロビーズ内に含有し，超音波を照射して徐放したのちに細胞に遺伝子導入ができるかどうかを96ウェルプレートを用いたin vitro試験により観察した．その結果，超音波によりAAVが徐放され，細胞に遺伝子導入できることを確認した．一方で，AAVの徐放を促すタングステン粒子の有無による遺伝子導入率に優位な差はなかった．これはナノ粒子と比較してAAVはハイドロゲルから徐放しやすいことが要因として考えられた．このため，AAVの漏れを防ぐために，ハイドロゲルマイクロビーズ自体のゲル化をより強固にすること，さらに本実験で使用したホモジナイザーよりも低出力のランジュバン型振動子を設計・製作し，これを用いた低出力超音波によるAAV照射実験をこれから実施する．

(2) パラボラ型振動子の製作
内視鏡による視野を確保しながら正円窓近傍でMHz帯の超音波を照射するため，小型で超音波の照射出力を高めることができる圧電素子にアルミ製のパラボラ板を取り付けたパラボラ型振動子を設計し，製作した．製作した超音波振動子は直径2.5 mmで，焦点距離2 mmのパラボラ板を0.5 mmの厚さの圧電素子に圧着して製作した．さらに，感電を防ぐために電極をエポキシ樹脂でシーリングすることで電極を被覆した．なお，製作したパラボラ型振動子は視野確保できる大きさを考えて，最大直径を3 mm以下とするように製作した．
In this year's experiments, the following two experiments were performed with the ultimate goal of gene transfer to the inner ear.

(1) Sustained release of AAV by ultrasonic irradiation
We contained AAV in hydrogel microbeads, irradiated them with ultrasound and released them slowly, and then observed whether gene transfer to cells could be achieved by in vitro testing using 96-well plates. As a result, it was confirmed that AAV was released slowly by ultrasound and that gene transfer to cells was possible. On the other hand, there was no significant difference in the gene transfer rate between the presence and absence of tungsten particles that promote the sustained release of AAV. This may be due to the fact that AAV is more easily released from the hydrogel than nanoparticles. In order to prevent leakage of AAV, we will make the gelation of the hydrogel microbeads themselves stronger, and design and fabricate a Langevin-type transducer with lower power than the homogenizer used in this experiment, and conduct AAV irradiation experiments using this transducer with low-power ultrasound.

(2) Fabrication of parabolic transducers
In order to irradiate MHz-band ultrasonic waves near the regular window while maintaining the endoscopic field of view, a parabolic transducer was designed and fabricated by attaching an aluminum parabolic plate to a piezoelectric element, which is compact and can increase the ultrasonic irradiation output. The ultrasonic transducer is 2.5 mm in diameter with a focal length of 2 mm. The parabolic plate is piezoelectrically bonded to a 0.5 mm thick piezoelectric element. In addition, the electrodes were covered with epoxy resin sealing to prevent electric shock. The parabolic transducer was fabricated so that its maximum diameter was less than 3 mm, considering the size of the field of view.