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Elastic fibres confer recoil and patency to the trachea and main bronchi, but systematic quantitative evidence for normal airway ageing is limited.
To quantify age-related changes in elastic fibre architecture of the human trachea and main bronchi.
In a cross-sectional cadaveric study (September 2025-January 2026), full-thickness tracheal and main bronchial segments were sampled from 41 adults (21-93 years) and stratified into four age groups (18-39, 40-59, 60-79, ≥80 years). Verhoeff-Van Gieson staining was used for elastin visualization. From four regions (tracheal anterior and posterior walls; right and left main bronchi), 775 quality-controlled fields (20x) underwent morphometry for elastic area fraction, fibre density, fragmentation index, and fibre thickness; observers were blinded to age. Non-parametric group comparisons and Spearman correlations assessed age effects.
Elastic area fraction declined progressively with age in all regions (all p<0.001), with reductions of ~29-41% from the youngest to oldest groups. Fibre density decreased by ~29-32% (all p<0.001). Fragmentation increased more than three-fold across regions (all p<0.001) and showed very strong positive correlations with age (r=0.893-0.947). Elastic area fraction correlated strongly and negatively with age (r=-0.785 to -0.850). Multivariable regression confirmed age as the dominant predictor, while sex effects were not significant. The posterior tracheal wall consistently retained higher elastic area fraction than the anterior wall (overall p<0.001).
Normal ageing is associated with substantial loss and fragmentation of airway elastic fibres, supporting observed declines in expiratory flow and cough effectiveness in healthy elders.
Keywords: Elastic fibres; Trachea; Main bronchi; Ageing; Verhoeff-Van Gieson; Morphometry.
Ethics approval: The study protocol received approval from the Institutional Ethics Committee.
Conflict of interest: None.
Author's contribution: All the authors equally contributed for the study and approved the final draft copy of the publication.
The elastic fibre network of the respiratory tract provides critical structural support and mechanical resilience to airways during cyclical breathing, coughing, and forced expiratory maneuvers. Composed of elastin protein embedded within a scaffold of fibrillin microfibrils, these fibres enable tissue recoil and maintain luminal patency against compressive forces. In the trachea and main bronchi, elastic fibres form dense lamellae within the submucosa and around cartilaginous plates, contributing to the biomechanical properties essential for efficient airflow [1,2].
Aging universally affects extracellular matrix components throughout the body. Elastic fibres undergo progressive structural deterioration characterized by fragmentation, thinning, reduced density, and disorganization of the fibrillar architecture. These changes reflect cumulative mechanical fatigue, enzymatic degradation, oxidative damage, and impaired synthesis of new elastin, which ceases after adolescence in humans. While age-related elastic fibre changes have been extensively documented in vascular tissues and pulmonary parenchyma, the conducting airways have received comparatively limited systematic investigation [3,4].
Clinical observations suggest functional consequences of airway aging. Healthy elderly individuals demonstrate altered respiratory mechanics including increased airway compliance, reduced peak expiratory flow rates, and modified cough effectiveness compared to younger adults, even without diagnosed pulmonary disease. Whether structural deterioration of elastic fibres underlies these physiological changes remains incompletely established through direct anatomical evidence [5,6].
Previous histological studies examining airway elastic fibres have been limited by small sample sizes, lack of age stratification, qualitative rather than quantitative assessments, or focus on diseased rather than normal aging. Precise morphometric quantification across standardized anatomical regions and well-defined age groups is needed to establish baseline patterns of structural senescence [7].
This study aimed to comprehensively evaluate age-related changes in elastic fibres of the human trachea and main bronchi using rigorous histological methods and quantitative morphometry. We hypothesized that elastic fibre density and integrity decline progressively with advancing age while fragmentation increases. By examining multiple anatomical regions across a broad age range with blinded semi-quantitative validation, we sought to provide robust evidence characterizing the structural basis for age-related alterations in large airway mechanics.
PK ej]7 OEBPS/s2-methods.xhtmlThis cross-sectional cadaveric histological study examined elastic fibre architecture in the trachea and main bronchi across four age groups. Specimens were obtained from routine dissection cadavers donated through institutional body donation programs during September 2025 to January 2026. The study protocol received approval from the Institutional Ethics Committee. Cadavers aged 18 years and above with intact airways were eligible for inclusion. Exclusions comprised specimens with gross airway pathology, prior tracheostomy, prolonged intubation, severe decomposition, or known chronic obstructive pulmonary disease when medical history was available. During standard anatomical dissection, the trachea and main bronchi were carefully exposed and harvested. A 2-cm mid-tracheal segment was excised to include both anterior cartilaginous and posterior membranous walls. Right and left main bronchus segments measuring 1 cm each were obtained at standardized distances from the carina. Specimens were rinsed in normal saline, assigned coded identifiers, and immersed immediately in 10% neutral buffered formalin at a tissue-to-fixative ratio of 1:10 for 24-48 hours.
Fixed specimens underwent routine tissue processing through graded ethanol dehydration, xylene clearing, and paraffin embedding. Blocks were oriented to obtain full-thickness cross-sections perpendicular to the luminal surface. Serial 4-5 µm sections were cut using a rotary microtome. Sections were stained with hematoxylin-eosin for morphological orientation and Verhoeff-Van Gieson stain as the primary method for elastic fibre visualization. The Verhoeff-Van Gieson technique renders elastic fibres black against a yellow background with red collagen, providing excellent contrast for morphometric analysis. Positive controls from elastic arteries accompanied each staining batch to ensure consistency [8,9].
A calibrated research microscope with attached digital camera captured standardized images at 20x magnification. Four regions of interest were defined per cadaver: tracheal anterior wall, tracheal posterior wall, right main bronchus, and left main bronchus. Within each region, five non-overlapping microscopic fields were systematically photographed from the submucosa, avoiding artifacts, folds, and edge effects. Images were saved as high-resolution files with standardized naming conventions linking them to specimen codes.
Quantitative analysis employed validated image processing software. For each field, elastic fibre area fraction was calculated by thresholding elastin-positive pixels and expressing as percentage of total region area. Fibre density quantified discrete fibres per square millimeter. Fragmentation index counted visible discontinuities where fibres exhibited gaps exceeding 5 micrometers, normalized to tissue area. Mean fibre thickness was measured across representative fibres using calibrated digital tools. Two independent observers, blinded to specimen age, assigned semi-quantitative scores from zero to three based on predefined criteria for fibre integrity [10].
Field-level measurements were averaged to cadaver-region level for analysis. Descriptive statistics included means and standard deviations. Normality was assessed using Shapiro-Wilk tests. Age-group comparisons employed Kruskal-Wallis tests with post-hoc pairwise testing and Bonferroni correction. Continuous age correlations used Spearman rank coefficients with bootstrap confidence intervals. Paired regional comparisons within cadavers utilized Wilcoxon signed-rank tests. Multiple linear regression models adjusted for age, sex, and anatomical region. Inter-observer agreement was quantified using Cohen's kappa. Statistical significance was defined as p < 0.05. All analyses were performed using standard statistical software.
PK ej]ҫX1@ @ OEBPS/s3-results.xhtmlForty-one cadavers were included in the final analysis after excluding 3 specimens due to tissue processing artifacts (n=2) or incomplete data collection (n=1). The study cohort comprised 28 males (68.3%) and 13 females (31.7%) with ages ranging from 21 to 93 years (mean 62.7 ± 20.6 years). Specimens were stratified into four age groups: 18-39 years (n=6, mean age 27.3 ± 3.6 years), 40-59 years (n=10, mean age 48.1 ± 3.7 years), 60-79 years (n=15, mean age 70.1 ± 2.8 years), and ≥80 years (n=10, mean age 87.5 ± 3.5 years). All specimens were obtained from routine dissection cadavers donated through institutional body donation programs. Fixation parameters were standardized across all age groups (mean fixation duration 37.1 ± 6.1 hours, range 30-50 hours) with no significant differences between age strata (p=0.961). A total of 880 microscopic fields (20x magnification) were captured from four anatomical regions per cadaver: tracheal anterior wall, tracheal posterior wall, right main bronchus, and left main bronchus. Of these, 775 fields (88.1%) passed quality control criteria and were included in morphometric analysis.
| Characteristic | 18-39 years | 40-59 years | 60-79 years | ≥80 years | Overall |
|---|---|---|---|---|---|
| Total Cadavers, n | 6 | 10 | 15 | 10 | 41 |
| Age (years), mean ± SD | 27.3 ± 3.6 | 48.1 ± 3.7 | 70.1 ± 2.8 | 87.5 ± 3.5 | 62.7 ± 20.6 |
| Age range | 21-32 | 42-53 | 64-74 | 83-93 | 21-93 |
| Male, n (%) | 3 (50.0) | 9 (90.0) | 7 (46.7) | 9 (90.0) | 28 (68.3) |
| Female, n (%) | 3 (50.0) | 1 (10.0) | 8 (53.3) | 1 (10.0) | 13 (31.7) |
| Fixation duration (hours), mean ± SD | 37.5 ± 5.5 | 36.8 ± 7.0 | 37.1 ± 5.8 | 37.2 ± 7.1 | 37.1 ± 6.1 |
| Total microscopic fields collected | - | - | - | - | 880 |
| Fields passing QC, n (%) | - | - | - | - | 775 (88.1) |
| Cadavers excluded | - | - | - | - | 3 |
Elastic area fraction showed highly significant progressive decline with age across all four anatomical regions (all p<0.001). In the tracheal anterior wall, elastic area fraction decreased from 17.97 ± 0.82% in the youngest group to 11.33 ± 1.88% in the oldest group (37% reduction). Similar patterns were observed in the tracheal posterior wall (22.42 ± 0.97% to 16.00 ± 1.94%, 29% reduction), right main bronchus (16.23 ± 1.06% to 10.05 ± 1.88%, 38% reduction), and left main bronchus (14.96 ± 0.90% to 8.77 ± 1.78%, 41% reduction). Fibre density decreased from 517.04 ± 47.73 to 348.98 ± 34.13 per mm2 in tracheal anterior wall (32% reduction) and ~29-30% in main bronchi (all p<0.001). The fragmentation index increased more than three-fold across all regions (tracheal anterior wall: 48.10 ± 11.00 to 149.93 ± 25.90 per mm2, p<0.001; right bronchus: 57.62 ± 11.33 to 172.32 ± 20.62 per mm2, p<0.001). Mean fibre thickness showed modest reductions (p<0.05 in anterior trachea, posterior trachea, and left bronchus).
| Region | Parameter | 18-39 years (n=6) | 40-59 years (n=10) | 60-79 years (n=15) | ≥80 years (n=10) | p-value |
|---|---|---|---|---|---|---|
| Trachea-Anterior | Elastic Area Fraction (%) | 17.97 ± 0.82 | 16.83 ± 1.24 | 14.46 ± 1.17 | 11.33 ± 1.88 | <0.001*** |
| Trachea-Anterior | Fiber Density (per mm2) | 517.04 ± 47.73 | 451.25 ± 38.44 | 424.96 ± 51.50 | 348.98 ± 34.13 | <0.001*** |
| Trachea-Anterior | Fragmentation Index (per mm2) | 48.10 ± 11.00 | 76.99 ± 12.04 | 117.07 ± 18.96 | 149.93 ± 25.90 | <0.001*** |
| Trachea-Anterior | Mean Fiber Thickness (µm) | 2.21 ± 0.16 | 2.00 ± 0.23 | 2.04 ± 0.34 | 1.68 ± 0.36 | 0.016* |
| Trachea-Posterior | Elastic Area Fraction (%) | 22.42 ± 0.97 | 21.59 ± 2.00 | 19.08 ± 1.09 | 16.00 ± 1.94 | <0.001*** |
| Trachea-Posterior | Fiber Density (per mm2) | 544.93 ± 38.04 | 507.33 ± 31.28 | 448.53 ± 42.03 | 405.47 ± 38.41 | <0.001*** |
| Trachea-Posterior | Fragmentation Index (per mm2) | 39.90 ± 7.05 | 69.51 ± 12.38 | 101.61 ± 14.20 | 147.98 ± 15.91 | <0.001*** |
| Trachea-Posterior | Mean Fiber Thickness (µm) | 2.24 ± 0.10 | 2.06 ± 0.20 | 2.07 ± 0.30 | 1.70 ± 0.36 | 0.019* |
| Right Bronchus | Elastic Area Fraction (%) | 16.23 ± 1.06 | 15.24 ± 1.79 | 13.06 ± 1.55 | 10.05 ± 1.88 | <0.001*** |
| Right Bronchus | Fiber Density (per mm2) | 470.40 ± 38.03 | 432.46 ± 32.89 | 393.07 ± 42.40 | 335.13 ± 29.68 | <0.001*** |
| Right Bronchus | Fragmentation Index (per mm2) | 57.62 ± 11.33 | 85.40 ± 18.49 | 121.30 ± 16.32 | 172.32 ± 20.62 | <0.001*** |
| Right Bronchus | Mean Fiber Thickness (µm) | 2.01 ± 0.07 | 1.82 ± 0.20 | 1.84 ± 0.36 | 1.52 ± 0.40 | 0.082 |
| Left Bronchus | Elastic Area Fraction (%) | 14.96 ± 0.90 | 15.04 ± 1.79 | 12.12 ± 1.33 | 8.77 ± 1.78 | <0.001*** |
| Left Bronchus | Fiber Density (per mm2) | 471.75 ± 48.96 | 439.88 ± 74.08 | 368.86 ± 37.25 | 333.60 ± 35.19 | <0.001*** |
| Left Bronchus | Fragmentation Index (per mm2) | 57.66 ± 6.09 | 84.58 ± 11.54 | 121.01 ± 18.45 | 161.66 ± 21.56 | <0.001*** |
| Left Bronchus | Mean Fiber Thickness (µm) | 2.01 ± 0.16 | 1.79 ± 0.23 | 1.90 ± 0.31 | 1.51 ± 0.42 | 0.028* |
| Region | Parameter | r (Spearman) | 95% CI | p-value |
|---|---|---|---|---|
| Trachea-Anterior | Elastic Area Fraction (%) | -0.850 | (-0.918, -0.720) | <0.001*** |
| Trachea-Anterior | Fiber Density (per mm2) | -0.794 | (-0.886, -0.618) | <0.001*** |
| Trachea-Anterior | Fragmentation Index (per mm2) | 0.893 | (0.781, 0.946) | <0.001*** |
| Trachea-Anterior | Mean Fiber Thickness (µm) | -0.454 | (-0.682, -0.144) | 0.003** |
| Trachea-Posterior | Elastic Area Fraction (%) | -0.812 | (-0.900, -0.673) | <0.001*** |
| Trachea-Posterior | Fiber Density (per mm2) | -0.815 | (-0.890, -0.688) | <0.001*** |
| Trachea-Posterior | Fragmentation Index (per mm2) | 0.941 | (0.861, 0.972) | <0.001*** |
| Trachea-Posterior | Mean Fiber Thickness (µm) | -0.463 | (-0.701, -0.155) | 0.002** |
| Right Bronchus | Elastic Area Fraction (%) | -0.785 | (-0.886, -0.597) | <0.001*** |
| Right Bronchus | Fiber Density (per mm2) | -0.744 | (-0.847, -0.559) | <0.001*** |
| Right Bronchus | Fragmentation Index (per mm2) | 0.923 | (0.830, 0.956) | <0.001*** |
| Right Bronchus | Mean Fiber Thickness (µm) | -0.372 | (-0.639, -0.058) | 0.017* |
| Left Bronchus | Elastic Area Fraction (%) | -0.833 | (-0.892, -0.705) | <0.001*** |
| Left Bronchus | Fiber Density (per mm2) | -0.695 | (-0.806, -0.498) | <0.001*** |
| Left Bronchus | Fragmentation Index (per mm2) | 0.947 | (0.868, 0.978) | <0.001*** |
| Left Bronchus | Mean Fiber Thickness (µm) | -0.375 | (-0.634, -0.034) | 0.016* |
| Comparison | Outcome | 18-39 years | 40-59 years | 60-79 years | ≥80 years | Overall |
|---|---|---|---|---|---|---|
| Trachea: Anterior vs Posterior | Elastic Area Fraction (%) | 0.031* | 0.002** | <0.001*** | 0.002** | <0.001*** |
| Trachea: Anterior vs Posterior | Fragmentation Index (per mm2) | 0.156 | 0.064 | 0.007** | 1.000 | 0.001** |
| Main Bronchi: Right vs Left | Elastic Area Fraction (%) | 0.031* | 0.375 | 0.095 | 0.006** | <0.001*** |
| Main Bronchi: Right vs Left | Fragmentation Index (per mm2) | 0.844 | 0.922 | 0.804 | 0.105 | 0.291 |
| Section / Metric | Value / Score 0 (%) | Score 1 (%) | Score 2 (%) | Score 3 (%) | Details / p-value |
|---|---|---|---|---|---|
| A. Inter-Observer Agreement (Overall, n=775) | Kappa = 0.217 (Fair) | 95% CI: 0.166-0.268 | - | - | Perfect Agreement: 43.2% |
| B. Age 18-39 years (n=116 fields) | 63.8% | 19.0% | 17.2% | 0.0% | χ² = 287.3, p<0.001 |
| B. Age 40-59 years (n=191 fields) | 46.1% | 22.0% | 31.9% | 0.0% | - |
| B. Age 60-79 years (n=288 fields) | 19.8% | 20.1% | 57.3% | 2.8% | - |
| B. Age ≥80 years (n=180 fields) | 3.9% | 3.9% | 74.4% | 17.8% | - |
| C. Mean Score: Trachea-Anterior | 0.55 ± 0.78 (18-39y) | 0.83 ± 0.87 (40-59y) | 1.33 ± 0.87 (60-79y) | 2.00 ± 0.42 (≥80y) | p<0.001*** |
| C. Mean Score: Trachea-Posterior | 0.07 ± 0.25 (18-39y) | 0.20 ± 0.50 (40-59y) | 0.92 ± 0.82 (60-79y) | 1.66 ± 0.78 (≥80y) | p<0.001*** |
| C. Mean Score: Right Main Bronchus | 0.79 ± 0.86 (18-39y) | 1.29 ± 0.85 (40-59y) | 1.74 ± 0.77 (60-79y) | 2.25 ± 0.44 (≥80y) | p<0.001*** |
| C. Mean Score: Left Main Bronchus | 0.75 ± 0.84 (18-39y) | 1.13 ± 0.82 (40-59y) | 1.74 ± 0.58 (60-79y) | 2.33 ± 0.52 (≥80y) | p<0.001*** |
| D. Correlation with Elastic Area Fraction | Spearman r = -0.645 | 95% CI: (-0.686, -0.603) | - | - | p<0.001*** |
| D. Correlation with Fragmentation Index | Spearman r = 0.612 | 95% CI: (0.564, 0.654) | - | - | p<0.001*** |
| Model / Outcome | Predictor | Beta Coefficient | 95% CI | p-value |
|---|---|---|---|---|
| Model 1: Elastic Area Fraction (%) (R2=0.847) | Age (per year) | -0.095 | (-0.108, -0.082) | <0.001*** |
| Model 1: Elastic Area Fraction (%) | Sex (Male vs Female) | -0.38 | (-1.12, 0.36) | 0.312 |
| Model 1: Elastic Area Fraction (%) | ROI: Trachea-Posterior | 4.52 | (3.81, 5.23) | <0.001*** |
| Model 1: Elastic Area Fraction (%) | ROI: Right Bronchus | -1.89 | (-2.60, -1.18) | <0.001*** |
| Model 1: Elastic Area Fraction (%) | ROI: Left Bronchus | -2.67 | (-3.38, -1.96) | <0.001*** |
| Model 2: Fragmentation Index (per mm2) (R2=0.881) | Age (per year) | 1.62 | (1.46, 1.78) | <0.001*** |
| Model 2: Fragmentation Index (per mm2) | Sex (Male vs Female) | 3.21 | (-3.15, 9.57) | 0.319 |
| Model 2: Fragmentation Index (per mm2) | ROI: Trachea-Posterior | -5.83 | (-12.01, 0.35) | 0.064 |
| Model 2: Fragmentation Index (per mm2) | ROI: Right Bronchus | 12.56 | (6.42, 18.70) | <0.001*** |
| Model 2: Fragmentation Index (per mm2) | ROI: Left Bronchus | 10.38 | (4.24, 16.52) | 0.001** |
This comprehensive histomorphometric study provides robust anatomical evidence for progressive age-related deterioration of elastic fibres in the human trachea and main bronchi. Analysis of 775 quality-controlled microscopic fields from 41 cadavers spanning seven decades revealed significant quantitative changes across all measured parameters. Elastic area fraction declined by 29-41% from young adulthood to advanced age, while fragmentation increased more than three-fold. These findings align with established concepts of extracellular matrix senescence but extend previous work by providing precise quantification across standardized anatomical regions with rigorous quality control [12,13].
The strong negative correlations between continuous age and elastic fibre integrity (r = -0.79 to -0.85) demonstrate that chronological aging dominates as the primary determinant of structural changes. This relationship persisted after controlling for sex and anatomical region in multivariate models, which explained over 84% of variance in elastic parameters [14,15]. The exponential increase in fragmentation compromises coordinated recoil properties and uniform force distribution. In large airways, this structural failure may contribute to increased compliance and reduced peak expiratory flows in elderly individuals [18,19].
Regional anatomical differences persisted across the lifespan, with the tracheal posterior membranous wall maintaining 20-25% higher elastic area fraction than the anterior cartilaginous wall, reflecting dynamic functional demands during cough and respiration. Paired main bronchi showed symmetric age-related changes without significant laterality in fragmentation (p = 0.291), supporting systemic ECM aging [20].
PK ej]6 OEBPS/s5-conclusion.xhtmlThis histomorphometric study demonstrates significant age-related deterioration of elastic fibres in the human trachea and main bronchi, with elastic area fraction declining 29-41% and fragmentation increasing three-fold from young adulthood to advanced age. Strong correlations confirm age as the dominant determinant, explaining over 84% of structural variance. Regional anatomical hierarchy persists throughout life, with the tracheal posterior wall maintaining superior preservation. These anatomical changes provide mechanistic insight into age-related alterations in airway compliance and respiratory mechanics, with implications for clinical assessment and surgical planning in elderly populations.
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eISSN 2321-4287 · pISSN 2321-8967
Volume 14 · Issue 2 · Pages 9511–9523 · June 2026
Research Article
1Department of Anatomy, Government Medical College, Paderu, ASR Dist, 531024, Andhra Pradesh, India.
2Department of Anatomy, PSP Medical College Hospital and Research Institute, Tambaram Kanchipuram Main Road, Oragadam, Panruti, Sriperumbudur Taluk, Kancheepurum Dist, Tamilnadu, 631604, India.
Corresponding author: Dr. Doni R Praveen Kumar, Assistant professor, Department of Anatomy, Government Medical College, Paderu, ASR Dist, 531024, Andhra Pradesh, India. E-Mail: donipraveen66@gmail.com
Received: 25 February 2026 · Revision received: 15 April 2026 · Accepted: 13 May 2026 · Published: 05 June 2026