UC San Diego

Skeletal muscle fibrosis is a hallmark of muscle injury and disease, and is defined as a general increase in extracellular matrix components including collagen. In addition to an increase in collagen, fibrotic muscle stiffness increases which manifests clinically as decreased range of motion and contracture. Despite the lack of clinical treatment for skeletal muscle fibrosis, very little data exist on extracellular matrix collagen and its alterations with fibrosis. This dissertation aims to elucidate the structure of extracellular collagen and its role in fibrosis. Because so little is known about the skeletal muscle extracellular matrix, Chapter 2 provides a review of the literature and original scanning electron micrographs of collagen structures that appear to be load- bearing. This review concluded that further study of the extracellular matrix was necessary to determine its normal structure and function before trying to interpret its fibrotic response. As a first attempt to quantify normal properties of the extracellular matrix, Chapter 3 describes a method for direct investigation of structure, composition, and mechanical properties. In this method, muscle fibers were removed from muscle without proteolytic enzymes or detergents that could degrade extracellular components. Passive mechanical testing was performed on intact extracellular matrix to directly measure its mechanical properties for the first time. With the observation of collagenous cable-like structures at the micron scale, a method was developed to better investigate extracellular collagen ultrastructure. Chapter 4 presents a three-dimensional electron microscopy method for reconstruction of skeletal muscle extracellular collagen structure. Collagen cable structures, fibroblasts, muscle fibers, and their interactions were modeled. With this new tool, it was possible to investigate the structure of fibrotic extracellular matrix collagen. Chapter 5 investigates the structure and quantity of collagen cables structures in normal and fibrotic muscle. Previous studies indicated that fibrotic muscle was stiffer than normal, so it was hypothesized that the volume fraction of collagen cables increased in fibrotic muscle. Fibrotic muscle contained double the volume fraction of collagen cables compared to wild type muscle. Collagen cables may therefore provide a future therapeutic target to reduce stiffness and improve function of fibrotic muscle