D.P.24 Identification of muscle necrosis in mdx mice using optical birefringence imaging

Abstract

Duchenne muscular dystrophy (DMD) is a severe form of muscular dystrophy, resulting in muscle degeneration and necrosis. Healthy skeletal muscle has a high degree of birefringence, caused by the regular arrangement of myofibrils. Birefringence decreases when the muscle becomes inflamed or necrotic and this arrangement breaks down. Hence birefringence is usable as a source of image contrast for identifying regions of damage. The mdx mouse is a well-established animal model for DMD. Assessment of the extent of muscle damage and necrosis in these mice is performed by analysis of histological sections, but this requires excision and fixation of the tissue. Optical coherence tomography (OCT) is an imaging modality which uses reflected near-infrared light to provide high resolution (≈10μm) images with a penetration depth of 1–2mm. Early studies on ex vivo tissue have shown OCT’s ability to differentiate between regions of intact myofibers and necrotic lesions. Other studies have demonstrated in vivo imaging deep below the surface by using OCT probes housed within hypodermic needles. Detection of birefringence is also possible using a variant known as polarisation-sensitive OCT (PS-OCT). In birefringent tissue such as muscle this can provide better contrast than regular OCT. In this study, we present a novel, automated, image analysis technique to quantify the birefringence present in a PS-OCT scan. The technique measures the average birefringence of muscle within an area 10×10μm across by 500μm deep. By translating this analysis across the tissue, we created a 2D image for each sample, representing variations in birefringence over the sample. Validated against H&E histology, the technique shows a strong correlation, with regions of low and high birefringence corresponding to areas of necrosis/inflammation and intact myofibers, respectively. This technique has the potential for in vivo assessment of muscle pathology without the need for staining or labelling.