Muscle Imaging Project Using ANatomopathology and Full Field Optical Coherence Tomography

Introduction Myositis represents a group of inflammatory diseases affecting skeletal muscles, characterized by inflammatory infiltration of muscle tissue. This condition can have various etiologies, such as infections, autoimmune diseases (like dermatomyositis or polymyositis), or drug-induced diseases. Early diagnosis and accurate assessment of muscle inflammation are essential for the therapeutic management of these patients.

Conventional analytical techniques, such as histopathology, allow visualization and analysis of muscle lesions and inflammation using stains on histological sections. However, this method requires several weeks of work before a result can be obtained. Full-field optical coherence tomography (OCT), a non-invasive imaging technique, could offer significant advantages in terms of speed of examination and resolution of muscle structures, particularly for visualizing inflammatory infiltration.

This research project proposes to compare conventional histopathology and full-field optical coherence tomography for the analysis of muscle biopsies from both healthy individuals and patients with suspected myositis.

The main objective of this prospective and comparative study will be to first establish a reference standard by analyzing control biopsies using both methods, and then to similarly evaluate the muscle inflammation detected in confirmed cases of myositis. The respective advantages and limitations of each technique will be assessed in the detection of this pathology.

Research Objectives

1. To analyze the spatial resolution of both methods for visualizing histopathological details at the level of muscle fibers, connective tissue, and blood vessels.
2. To compare the ability of conventional histopathology and full-field optical coherence tomography (OCT) to detect muscle inflammation in muscle biopsies from healthy individuals and patients with myositis.
3. To evaluate the diagnostic accuracy of both techniques in identifying inflammation-related abnormalities, such as inflammatory cell infiltration (lymphocytes, macrophages), muscle fiber degradation, and fibrosis.
4. To compare the acquisition time and ease of use of each method for analyzing muscle tissue in a clinical setting.

Hypotheses Full-field optical coherence tomography will enable real-time visualization of inflammatory infiltration and muscle abnormalities with microscopic resolution, in a non-invasive, rapid, and dynamic manner.

These results could very quickly complement those obtained by anatomopathology which aims to describe the classic histopathological aspects of myositis, such as muscle necrosis, inflammatory infiltration, and fibrosis.

Methodology

1. Sample Selection

• Patient Groups:
• Group 1 (healthy patients): Muscle biopsies from approximately ten healthy adult patients with no history of muscular, neurodegenerative, or inflammatory disease, undergoing orthopedic surgery. The age range should be variable (18-90 years) to be representative of the population and to account for natural age-related atrophy.
• Group 2 (patients with suspected myositis): Muscle biopsies from adult patients presenting with clinical symptoms consistent with myositis (muscle fatigue, pain, muscle weakness) and clinical suspicion confirmed by other diagnostic tests (e.g., serology, EMG, clinical examination). Healthy muscle tissue will be obtained from a resection necessary during orthopedic surgery. This surgical waste does not constitute additional samples. Biopsies for suspected myositis will be performed as part of the diagnostic process and do not require any additional sampling. An information sheet will be given to the patient, and the surgeon will obtain their consent before the procedure. All patients must be of legal age, affiliated with or covered by the social security system, and not subject to any legal guardianship or protective measures. Pregnant women and adults unable to express their consent cannot be included in the research protocol.

1. Conventional Anatomopathological Analysis

• Samples will be frozen in ispoentane, stored at -80°C, and then transported on dry ice to the Anatomopathology Department of the Hospices Civils de Lyon for section labeling.
• Histological staining of healthy tissues: Three standard stains will be performed on control biopsies (hematoxylin and eosin (H\&E), Gomori stain, fiber typing) to collect structural and metabolic data.
• Histological staining in cases of suspected myositis: The three standard stains will also be performed, as well as stains for inflammatory markers, namely the major histocompatibility complex (HLA I, HLA DR), complement (C5b9), autophagy (p62), and clusters of differentiation (CD56, CD31).

1. Full-field optical coherence tomography (OCT) analysis

• Full-field OCT technology allows for micrometer-resolution imaging without the need for prior sample preparation. OCT will be used to image muscle tissue samples in real time, both structurally (FF-OCT) and dynamically (DCI). A series of 1 µm slices will be acquired. The images will then be converted by an algorithm to reproduce hematoxylin and eosin staining.
• OCT images could allow visualization of muscle fiber structure, surrounding connective tissue, and areas of inflammatory infiltration.

1. Comparison of methods Qualitative analysis: The images obtained by OCT will be visually compared to images obtained by conventional histology after scanning the slides. This comparison will be conducted by specialized anatomical pathologists to evaluate the ability of each method to detect muscle abnormalities and inflammation. An artificial intelligence algorithm may be used to facilitate the comparison.
2. Expected Results The results will allow us to evaluate the clinical value of OCT as an alternative or complementary imaging method to conventional histopathology. If OCT proves effective for detecting muscle inflammation, it could become a valuable tool for ultra-rapid diagnosis.