Musculoskeletal injury and repair

Bone repair, whether it happens following a fracture or a bone graft, involves a well organized set of events that lead to reconstitution of the biological and mechanical integrity of bone. The regeneration process is initiated by an inflammatory response, which plays an important role in stimulating repair.¹ Simultaneously, skeletal progenitor cells are recruited and begin differentiating into chondrocytes and osteoblasts that will deposit new cartilage and bone matrix necessary for bone bridging. The origins of these progenitor cells and the influence of the inflammatory response on their recruitment are not well understood. Following extracellular matrix deposition, cartilage is replaced by bone and new trabecular bone is converted to lamellar bone during the remodeling phase of repair.^2–4

Numerous molecules and growth factors are keys to each step of the repair process and their functions are slowly being elucidated through analyses of various animal models of bone repair.^2,5 These investigations will lead to a better understanding of the cellular and molecular bases of bone repair, better diagnosis of skeletal repair defects, and development of new strategies to accelerate healing. Surgeons have now the choice between various surgical techniques, improved implants and biological approaches to treat complex injuries. Current approaches use autografts, allografts, or bone morphogenetic proteins (BMPs). However, these approaches are not always successful and are costly, which necessitates the development of new therapies.

The muscles, tendons and ligaments along with blood vessels and nerves are closely associated with the bone. Musculoskeletal injuries may involve one or more of these tissues and the extent of injury is highly linked to the success of repair. For example, delayed union or non-union occurs in 5 to 10% of all fractures but is increased up to 46% in patients with extreme trauma and soft tissue damage.⁶ Therefore the role of numerous tissues must be taken in account in the majority of musculoskeletal diseases or injuries. Advances are being made in the basic biology of bone and individual soft tissues surrounding bone. The basic biology of muscle and muscle repair is well understood compared to other soft tissues. Muscle repair is composed of three phases including degeneration/ inflammation, regeneration and fibrosis. Many molecular markers and disease models are available. Muscle has been an ideal target to test new gene therapies and cell based therapies, however further advances are needed to treat devastating diseases such as Duchenne Muscular Dystrophy and to improve muscle repair. Vascular biology is also an area of intense investigation but more efforts need to be made to apply the data to the orthopedic field. The biology of tendons and ligaments is now being better understood with the identification of key molecular pathways involved in these tissues. Injury of tendons and/or ligament independent of bone can lead to complications and extended periods of recovery, which can also have debilitating effects. Like muscle and bone healing, tendon and ligament healing is initiated by an inflammatory response that may be modulated to stimulate repair. Little is know about the intrinsic capacities of tendon and ligament to heal and the cell sources that participate in repair.

Cartilage biology

Regeneration of cartilage is a hot topic. Cartilage can be damaged by injury, inflammation, infection, and degeneration. Destruction of articular cartilage in rheumatoid arthritis and osteoarthritis involves inflammatory cytokines such as TNF-α, IL-1, or IL-6, which have been the target for current therapies.^7–11 After injury, articular cartilage has a poor capacity to repair itself and tends to heal through the formation of fibrocartilage, which has inferior biomechanical characteristics to resist compression stress compared to normal articular hyaline cartilage. For decades scientists and surgeons have been exploring treatments that facilitate cartilage repair, including micro fractures and more recently cell-based tissue engineering using autologous chondrocytes and mesenchymal stem cells combined with scaffolds. Mesenchymal stem cells can be collected from many sources including bone marrow, adipose tissue, synovium, and muscles. Study on embryonic stem cells has also been initiated and this research direction could be very fruitful. Many cell factors may be exploited to improve these approaches, among which are TGF-β, IGF-1, FGF-2, and BMP-7.¹² Nanotechonology may also improve the biomaterial properties of scaffolds. A better understanding of the cellular and molecular mechanisms of chondrocyte differentiation and phenotype maintenance may provide insights to develop novel techniques that can guide cultured chondrocytes and stem cells differentiation.

Regeneration of intervertebral discs (IVD) is equally, if not more challenging. A clinical study has shown that injection of disc chondrocytes reduces back pain and improves fluid contents of the treated disc.¹³ Experimental studies have demonstrated that injected mesenchymal stem cells can maintain viability and proliferate within the IVD.¹⁴ Although there are significant advances in the research of IVD regeneration, this area of research is still in its early stages. The intervertebral disc is composed of three tissues, the cartilage endplate, nucleus pulposus, and annulus fibrosus, and is more complex than articular cartilage. Successful regeneration of IVD may require the regeneration of all three tissues in one implant. The right scaffolds, cells, and techniques of implantation need to be determined and developed.¹⁵

Infrastructure and equipments: Molecular biology laboratory

The infrastructure required to run an Orthopaedic Surgery Research laboratory is similar to any other biological laboratory. Fume hoods are required to vent noxious and dangerous chemicals. An animal housing facility is necessary if work is performed on any number of model organisms. If work is to be performed on established or primary cell lines, then a separate cell culture room should be considered. By isolating cell culture facilities, reduced foot traffic in and around the incubators and hoods will aid in keeping cultures free of bacteria and mold. Another part of the laboratory should be set aside for processing, sectioning, and staining of histological specimens. This area should be located in a “dust-free” area away from drafts that will create difficulty handling ribbons of sections. Work with radioactive materials can be made safer by defining and restricting use of these materials to dedicated areas of the laboratory. Similarly a dedicated imaging suite that contains all the microscopes that will be used for documentation and analysis of data will allow undisturbed specimen viewing, will allow the room to be darkened for specialized imaging such as epifluorescence, and will reduce the amount of dust that accumulates on working parts of the microscope.

Equipment for an Orthopedic Surgery laboratory performing molecular and cellular biology experiments includes microtomes, thermal cyclers, bacterial shaking incubators and incubator ovens, electrophoresis equipment for assessing DNA, RNA, and proteins, table top microfuge, centrifuge, safety cabinets to store flammable liquids. If work is primarily focused on in vitro analyses, then cell culture incubator(s), laminar flow hoods, and at least one inexpensive inverted phase contrast microscope for visualizing cells are required. More specialized equipment can be used as the necessity of the laboratory dictates. For example, quantitative reverse transcriptase PCR (qPCR) can be used to assess gene expression patterns in cells and tissues.