拓宽 ARMC2 突变表型：将鞭毛的多种形态异常与原发性睫状肌运动障碍的肺部表现联系起来

Broadening the ARMC2 mutational phenotype: linking multiple morphological abnormalities of the Flagella to Pulmonary Manifestations in Primary Ciliary Dyskinesia

拓宽 ARMC2 突变表型：将鞭毛的多种形态异常与原发性睫状肌运动障碍的肺部表现联系起来

Authors: Baoyan Wu, Wenhao Zhang, Hui Yu, Lewen Ruan, Kai Wang, Meng Gu, Hao Geng, Jiajun Fang, Chuan Xu, Yuying Sheng, Qing Tan, Qunshan Shen, Zongliu Duan, Huan Wu, Rong Hua, Rui Guo, Zhaolian Wei, Ping Zhou, Yuping Xu, Yunxia Cao, Xiaojin He, Kuokuo Li, Mingrong Lv, Dongdong Tang

Source:Reproductive Biology And Endocrinology

DOI: 10.1186/s12958-025-01385-0

Abstract

Background: Severe asthenoteratozoospermia, a prevalent cause of male infertility, has increasingly been associated with ARMC2 variants that cause Multiple Morphological Abnormalities of the Sperm Flagella (MMAF). Although ARMC2 is also expressed in other ciliary structures, no studies have yet reported a link between ARMC2 gene variants and other symptoms of Primary Ciliary Dyskinesia (PCD).

Methods: Here, we performed whole-exome sequencing (WES) on Chinese subjects with MMAF to identify potential genetic variants. Sanger sequencing was used to validate the candidate variants. Sperm morphology was assessed using modified hematoxylin and eosin (H&E) staining, and transmission electron microscopy (TEM) was performed to observe the ultrastructural defects of the sperm flagella. Western blot analysis and immunofluorescence (IF) of spermatozoa were performed to evaluate variations in structural protein. Additionally, intracytoplasmic sperm injection (ICSI) was applied for assisted fertilization.

Results: We identified two compound heterozygous ARMC2 variants and one homozygous variant (P1: c.1030_1042del, p.T344fs/c.1331G > A, p.R444H; P2:c.1264C > T, p.R422X) in two unrelated individuals. Notably, in addition to MMAF, individual P2 exhibited classic symptoms of PCD in the lungs, including recurrent airway infections, bronchitis, and rhinosinusitis. Morphological and ultrastructural analyses of the spermatozoa obtained from the two individuals revealed dramatic disorganization in axonemal and peri-axonemal structures, as well as the absence of the axonemal central pair complex (CPC). Immunoblotting and immunofluorescence assays revealed the reduced expression of ARMC2 and the abnormality of various axonal structural proteins. Further assisted reproduction outcomes showed that one of the individuals conceived successfully after Intracytoplasmic Sperm Injection (ICSI).

Conclusions: Overall, this study significantly expanded the mutational phenotype of ARMC2, marking the first discovery of PCD-related pulmonary phenotypes outside of the reproductive system. This work establishes the association between ARMC2 and typical PCD and lays the groundwork for further investigation into the molecular mechanisms of ARMC2 in both flagellogenesis and ciliogenesis.

Keywords: ARMC2; Cilla; Flagellum; MMAF; Male infertility; PCD.

摘要

背景：严重的无精子症是导致男性不育的一个主要原因，越来越多的人认为该病与导致精子鞭毛多形态异常（MMAF）的 ARMC2 基因变异有关。虽然 ARMC2 也在其他睫状肌结构中表达，但目前还没有研究报道 ARMC2 基因变异与原发性睫状肌运动障碍（PCD）的其他症状之间存在联系。

方法：在此，我们对中国的 MMAF 患者进行了全外显子组测序（WES），以确定潜在的基因变异。桑格测序用于验证候选变异。采用改良苏木精和伊红（H&E）染色法评估精子形态，并通过透射电子显微镜（TEM）观察精子鞭毛的超微结构缺陷。对精子进行了 Western 印迹分析和免疫荧光（IF），以评估结构蛋白的变化。此外，还采用了卵胞浆内单精子显微注射（ICSI）技术进行辅助受精。

结果：我们在两个无血缘关系的个体中发现了两个复合杂合ARMC2变异体和一个同源变异体（P1：c.1030_1042del, p.T344fs/c.1331G > A, p.R444H；P2:c.1264C > T, p.R422X）。值得注意的是，除了 MMAF 外，个体 P2 还表现出肺部 PCD 的典型症状，包括反复气道感染、支气管炎和鼻炎。对这两个个体的精子进行的形态学和超微结构分析表明，轴突和轴突周围结构严重紊乱，轴突中央对复合体（CPC）缺失。免疫印迹和免疫荧光检测显示，ARMC2表达减少，各种轴突结构蛋白异常。进一步的辅助生殖结果显示，其中一人在卵胞浆内单精子显微注射（ICSI）后成功受孕。

结论：总之，这项研究大大扩展了 ARMC2 的突变表型，标志着首次在生殖系统之外发现了 PCD 相关的肺表型。这项工作确立了ARMC2与典型PCD之间的关联，并为进一步研究ARMC2在鞭毛发生和纤毛发生中的分子机制奠定了基础。

关键词： ARMC2；纤毛；鞭毛；MMAF；男性不育；PCD