Myofibre Density Reveals a Critical Threshold Around Age 6 in Steroid-Naïve Duchenne Muscular Dystrophy: A Retrospective Observational Study.

Abstract

Aims: In Duchenne muscular dystrophy (DMD), robust histological markers for assessing early disease progression remain elusive. We defined myofibre density (MFD) as the count of myofibres per square millimetre, which, in our preliminary survey of DMD muscle biopsies, steeply declined with age (Spearman's ρ: -0.85). We aimed to characterise age-dependent MFD dynamics in early-stage DMD.

Methods: We retrospectively assessed 46 archival muscle-biopsy slides (steroid-naïve; collected > 40 years ago) using semi-quantitative image analysis with digital restoration. MFD and classical histological variables were quantified. Age-MFD dynamics were modelled with segmented regression and validated using weakly informative Bayesian modelling. The primary measure was the MFD age-breakpoint. Secondary measures included breakpoint-detection power, age-predictive MFD cut-offs and behaviour of conventional variables across breakpoint-defined age bands.

Results: After quality and age-distribution screening, 35 slides (age 1-11 years) were analysed. Segmented regression identified a breakpoint at 6.25 years (95% confidence interval [CI]: 5.08-7.42); after which MFD plateaued at lower levels. Bayesian posterior estimate was 6.37 years (95% credible interval: 5.24-7.66). A 10,000-run Monte Carlo simulation (n = 35) showed approximately 80% power to recapture the breakpoint within ±1.25 years. MFD cut-offs > 596 and < 426 fibres/mm² corresponded to 80% and 20% probabilities of age < 6.25 years. Between the early (< 5 years) and late (≥ 7.5 years) age bands defined by the breakpoint-CI limits, myofibre-size parameters, myofibre area and fat replacement shifted significantly.

Conclusions: MFD, a simple metric, reveals a previously unrecognised phase of rapid myofibre loss lasting up to around age 6 in early-stage DMD.