利用腺相关病毒AAV1探究顺铂诱导的血管纹损伤中GASDERMIN D的作用机制

Investigating the mechanism of GASDERMIN D in cisplatin-induced stria vascularis injury using adeno-associated virus AAV1

摘要

目的 探究含有细胞焦亡关键执行蛋白GASDERMIN D(GSDMD)的基因片段及绿色荧光蛋白(Green fluorescent protein, GFP)标记的AAV1-GSDMD-GFP病毒对进行顺铂处理后的Gsdmd敲除小鼠(Gsdmd^-/-)模型血管纹(Stria vascularis, SV)形态学影响。方法 将Gsdmd^-/-小鼠分为四组，分别为对照组(Gsdmd^-/-)、空白对照组(Gsdmd^-/-+AAV1-GFP)、顺铂处理组(Gsdmd^-/-+Cisplatin)和回补GSDMD的顺铂处理组(Gsdmd^-/-+AAV1-GSDMD-GFP+Cisplatin)，通过荧光染色(Phalloidin、KCNQ1和GS-IB₄)来分析小鼠血管纹形态变化。结果 Gsdmd^-/-+Cisplatin组小鼠可以抵抗顺铂耳毒性，成功回补GSDMD后，血管纹出现形态学上的异常(血管树突状断裂、细胞形态异常)，表明GSDMD回补逆转了Gsdmd^-/-小鼠的抵抗性。结论 通过AAV1回补GSDMD可以逆转Gsdmd^-/-小鼠血管纹对顺铂耳毒性的抵抗。

Abstract

Objective To investigate the morphological effects of AAV1-GSDMD-GFP virus, containing the gene fragment of the key pyroptosis execution protein GASDERMIN D (GSDMD) labeled with green fluorescent protein (GFP), on the stria vascularis (SV) of Gsdmd knockout (Gsdmd^-/-) mice after cisplatin treatment. Methods Gsdmd^-/- mice were divided into four groups: Control (Gsdmd^-/-), blank virus control (Gsdmd^-/-+AAV1-GFP), cisplatin-treated (Gsdmd^-/-+Cisplatin), and GSDMD-rescued (Gsdmd^-/-+AAV1-GSDMD-GFP+Cisplatin). Morphological changes in the SV were analyzed via fluorescence staining (Phalloidin, Kcnq1, and GS-IB₄). Results The Gsdmd^-/-+Cisplatin group exhibited resistance to cisplatin-induced ototoxicity. However, successful reintroduction of GSDMD via AAV1-GSDMD-GFP resulted in significant morphological abnormalities in the SV, including vascular dendritic fragmentation and cellular structural disruption, indicating that GSDMD restoration reversed the resistance phenotype. Conclusion Resistance to cisplatin ototoxicity in the stria vascularis of Gsdmd^-/- mice can be reversed by AAV1 backfilling of GSDMD.

关键词

腺相关病毒 / 细胞焦亡 / 顺铂耳毒性 / 血管纹损伤

Key words

Adeno-associated virus / Pyroptosis / Cisplatin ototoxicity / Stria Vascularis Injury

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