Association of reduced DTI-ALPS index and increased free water with glymphatic system alterations in noise-induced hearing loss: a neuroimaging study

Subjects

According to the national standard of the People’s Republic of China, GBZ 49-2014 “Diagnosis of Occupational Noise Deafness”, 48 NIHL patients were recruited as research subjects from the occupational department of Yantaishan Hospital between 2014 and 2020. The NIHL group consisted predominantly of individuals engaged in rock drilling and welding. As a result, all participants were adult males. Meanwhile, 40 healthy controls (HCs) matched to the NIHL group for age, education level, and gender were enrolled in the study. The clinical characteristics of all the participants, including age, education level, better-ear monaural threshold weighted value (MTWV), and Hamilton Anxiety Scale (HAMA) score, were collected and analyzed. The diagnosing criteria for all participants: for individuals with an average hearing threshold above 40 dB in both ears for high-frequency ranges (3000, 4000, and 6000 Hz), diagnosis and classification are based on the weighted values of the better whispered frequency (500, 1000, and 2000 Hz) and the hearing threshold at 4000 Hz. NIHL is diagnosed when MTWV is ≥ 26 dB; while MTWV below 25 dB was diagnosed as normal23. All patients met the diagnostic criterion of MTWV ≥ 26 dB HL. All HCs had normal hearing defined as MTWV < 25 dB HL. By study design, only the binary inclusion criterion (normal hearing: yes/no) was recorded at the time of recruitment; exact MTWV values are not available for the HCs. The inclusion and exclusion criteria for all participants: adult Han Chinese males (due to the male-dominated nature of the occupations involved); educational level: primary school or above; participants had no personal/family history of psychiatric or neurological disorders and did not use psychotropic drugs.

The study was approved by the Ethics Committee of Yantaishan Hospital (Ethics Approval No.: Yanshanlun 2023014). All participants provided written informed consent. The entire study was conducted in strict accordance with the Declaration of Helsinki and relevant medical ethical norms to ensure the rights and interests of participants as well as data security.

Imaging acquisition and preprocessingImaging acquisition

The GE Discovery MR 750 3.0T with 8-channel brain coil was used for scanning all participants. The DTI sequence parameters included: TR = 5500ms, minimum TE, slice thickness = 3.0 mm, gap = 0 mm, FOV = 24 cm × 24 cm, flip angle = 90°, matrix size = 128 × 128, NEX = 1, b-value = 0,1000s/mm², gradient direction = 50, scanning time = 4 min 46 s.

Imaging preprocessing

Diffusion MRI data were processed with FMRIB Software Library (FSL, http://www.fmrib.ox.ac.uk/fsl/) and MRtrix3 (https://www.mrtrix.org/). The preprocessing steps included denoising, Gibbs artifact correction, eddy current and motion correction, B1 field bias correction, and skull stripping.

DTI-ALPS index calculation

The DTI-ALPS index was calculated with a validated semi-automated approach through bash scripts10,25. The procedure involved the following steps:

(1)

ROI definition. Spherical ROIs (5 mm diameter) were placed at the level of the lateral ventricles on projection and association fibers using the JHU-ICBM-FA template. The central coordinates of ROIs for bilateral projection fibers and association fibers were as follows: L_proj (116, 110, 99), L_assoc (128, 110, 99), R_proj (64, 110, 99), and R_assoc (51, 110, 99).

(2)

Images processing and registration. FA and tensor maps were generated with FSL. Individual FA maps were registered to the JHU-ICBM-FA template. The resulting transformation matrix was subsequently applied to align the corresponding tensor map to the template space. All registered images were manually inspected to ensure quality.

(3)

Diffusion values extraction. The diffusion components (Dxx, Dyy, Dzz) were extracted from the registered tensor maps, and their diffusion values were obtained within the predefined ROIs using the following variable conventions:

Dxxproj_L, Dxxproj_R: X-axis diffusivity in left/right projection fibers.

Dxxassoc_L, Dxxassoc_R: X-axis diffusivity in left/right association fibers.

Dyyproj_L, Dyyproj_R: Y-axis diffusivity in left/right projection fibers.

Dzzassoc_L, Dzzassoc_R: Z-axis diffusivity in left/right association fibers.

(4) DTI-ALPS index calculation. The left, right, and mean DTI-ALPS indices were calculated in the following manner:

Left DTI-ALPS = ((Dxxproj_L + Dxxassoc_L)/2) / ((Dyyproj_L + Dzzassoc_L)/2).

Right DTI-ALPS = ((Dxxproj_R + Dxxassoc_R)/2) / ((Dyyproj_R + Dzzassoc_R)/2).

Mean DTI-ALPS = ((Dxxproj_L + Dxxassoc_L + Dxxproj_R + Dxxassoc_R)/4) / ((Dyyproj_L + Dzzassoc_L + Dyyproj_R + Dzzassoc_R)/4).

FW in white matter calculation

As described in previous studies10,26, the regularized bi-tensor model was used to construct FW maps, and Tract-Based Spatial Statistics (TBSS)27was employed to obtain FW values along the white matter skeleton. The workflow was as follows:

(1)

Generate free water maps by fitting a regularized bi-tensor model using the Diffusion Imaging (Dipy) in Python open-source package28.

(2)

Using TBSS, create the group-level mean FA map from all subjects first, then extract FA skeleton map with an FA threshold level of 0.2.

(3)

Project the FW map of each participant onto the mean FA skeleton map to obtain its FW map on the white matter skeleton.

(4)

Extract FW values based on the skeletonized FW maps.

Statistical analysis

Statistical analyses were performed in R software (version 4.3.3; https://www.r-project.org/). The normality of continuous variables was evaluated via Shapiro-Wilk test. Independent sample t-tests or Mann-Whitney U tests were used to analyze group differences in demographic and clinical characteristics.

After controlling for age and education level, analysis of covariance (ANCOVA) was used to examine between-group differences in mean DTI-ALPS, left DTI-ALPS, right DTI-ALPS, and FW. Prior to the ANCOVA, the model assumptions were tested for normality of residuals and homoscedasticity using the Shapiro-Wilk test and Levene’s test, respectively. Both assumptions were met. Paired t-tests were applied to assess within-group differences in bilateral hemispheric DTI-ALPS within both the NIHL group and the HCs group.

Spearman correlation analysis was employed to explore the correlations between DTI-ALPS and clinical characteristics as well as FW. A value of p < 0.05 was considered statistically significant and Benjamini–Hochberg FDR correction was used for multiple comparisons.