Regenerative medicine (RM) encompasses an emerging field of medicine with the goal of replacing, engineering, or regenerating human cells, tissues, or organs lost or injured due to age, disease, or congenital defects to restore or establish normal function. Researcher scientists and medical practitioners are attempting to create new interventions to treat a variety of diseases that address every system in the body and treat chronic diseases such as diabetes, stroke, macular degeneration, congestive heart failure, and osteoarthritis. This chapter discusses the use of RM in the treatment of acute and chronic pain from painful musculoskeletal conditions.
RM therapies in the treatment of musculoskeletal conditions focus on promoting the body’s innate healing capacity. Interventions include using prolotherapy principles to inject concentrated dextrose, platelet-rich plasma (PRP) injections, and adipose or bone marrow-derived stem cells. Significant evidence exists for the use of RM in the treatment of osteoarthritis, tendinopathy, and ligamentous pathology. Importantly, the use of RM techniques is a shift in the treatment away from conventional destructive techniques such as corticosteroid and local anesthetic injections, and neurolysis. Numerous studies support that local anesthetic, corticosteroids, and contrast agents have deleterious effects on soft tissues including chondrocytes and tenocytes, the cells that constitute cartilage and tendons.
While RM has capacity for healing various musculoskeletal disorders, it is important to recognize the limitations of these modalities and select patients and pathology that will best respond to these various techniques. Although RM is generally most effective for mild-to-moderate disease, some studies have demonstrated success with more severe disease states. , , , Even so, surgical management or alternative ablative techniques may be more appropriate for complete tears or end-stage, grade 4 osteoarthritis, and thus, preprocedural patient selection is key to intervention success. Additional patient characteristics that would impair the body’s ability to heal or degrade its regenerative capacity include smoking cigarettes, uncontrolled blood glucose, immunosuppressed states, or active infections. Areas that lack adequate blood supply, such as eschars, avascular or necrotic limbs, are also unlikely to respond to RM techniques given their poor capacity to receive and utilize the necessary elements for healing.
The Healing Cascade
The healing cascade lays the framework for RM and occurs over the course of weeks, with final tissue remodeling taking potentially many months before restoration of full tissue strength and integrity. Healing involves many of the same activating signals and growth factors released by platelets during degranulation. The process of wound healing can be subdivided into inflammation, proliferation, and maturation stages.
Differing models of the cascade may separate out the process of hemostasis (coagulation) as occurring to prior to the inflammation stage, while others include it. Despite the taxonomical variance, the entire process occurs as part of the spectrum of healing. After the formation of a fibrin mesh and a clot, cytokines and growth factors previously released during platelet degranulation stimulate the complement cascade and recruit leukocytes (primarily neutrophils), macrophages, and fibroblasts to the injured area. Local histamine release leads to increased capillary permeability via vasodilation and leakage, allowing migration of mesenchymal stem cells (MSCs) to the site. Neutrophils then lead the process of decontamination through bacterial lysis and scavenging of cellular debris. Monocytes previously activated by platelet growth factors also migrate to the area and may differentiate into macrophages. These macrophages play various important roles: bacterial phagocytosis, cytokine and collagenase secretion for tissue remodeling, and secretion of growth factors that contribute toward angiogenesis and formation of granulation tissue. Among the factors secreted by macrophages are many that are associated with bone repair, such as interleukins (ILs), tumor necrosis factor alpha (TNF-α), transforming growth factor beta (TGF-β), platelet-derived growth factor (PDGF), epidermal growth factor (EGF), and vascular endothelial growth factor (VEGF). ,
The proliferation stage then begins with epithelialization by migratory epithelial progenitor cells as well as epithelial cells from the wound periphery. Angiogenesis takes place under the signals from previously released platelet growth factors. Fibroblasts drive the production of granulation tissue and collagen deposition around 4 days after an injury. MSCs are integral in coordinating the healing response, but can also be activated to begin differentiation down chondrogenic, osteogenic, or angiogenic pathways.
During the final maturation or remodeling stage, a wound contract as collagen continues to be deposited by fibroblasts, granulation tissue compresses into smaller and newly formed scar tissue, and the strength of this new wound increases. This process is also driven by various growth factors that were present for the previous steps. Overall, while it is easier to comprehend all these steps linearly, in reality many of them overlap and occur simultaneously providing an onslaught of regeneration and remodeling.
Prolotherapy
Proliferant solutions vary in the mechanism by which they cause localized inflammation but, in general, they all act by causing localized tissue trauma or irritation which initiates an inflammatory response. Various injectates include osmotics (dextrose, glycerin), chemotactics (sodium morrhuate) neurolytics (phenol, dextrose), local irritants, and particulates (guaiacol, tannic acid, pumice), with the most commonly used solution being dextrose 15%–25%.
Dextrose 15%–25% is a hyperosmolar shock agent, which results in shrinking of the cell, lipid leakage, thus inflammation. Some cell death also occurs. This leads to the release of various cellular parts, proteins, and membrane fragments which are attractive for granulocytes and macrophages, leading to stimulation of the healing cascade. Furthermore, the injection itself—especially with the “peppering technique”—causes tissue trauma, platelets activation, with release of bioactive cytokines and growth factors involved in the healing process.
The treatment regimen typically includes a series of injections to treat one injury. Many of the injection targets were traditionally described and injected based on landmark-guided technique. Some superficial structures-like supraspinous ligaments are still best targeted based on palpation; however, with the availability imaging, deeper anatomical structures are also precisely and safely injected with US or fluoroscopy guidance.
Platelet-Rich Plasma
PRP is an autologous blood product defined as a volume of plasma that has a supraphysiologic platelet count. These platelets contain over 30 biologically active growth factors stored in alpha granules. Thus, increased platelet count is an ad hoc measurement of growth factor concentration which can be delivered to damaged tissues to promote healing. Growth factors include VEGF, PDGF, TGF, platelet-derived angiogenesis factor, epidermal growth factor (EGF), fibroblast growth factor, and connective tissue growth factor. PRP has been used for various indications including optimizing orthopedic and spine surgical outcomes, dentistry, wound care, tightening of the skin, and even hair growth. Factors that are important in the PRP extraction process include purity (reducing the red blood cells and white blood cells), concentration factor, and total amount of blood collected. The total amount of blood collected will directly correlate with the resultant total amount of platelets in your PRP fraction.
Recent studies have demonstrated that higher platelet concentrations are ideal to promote healing of soft tissue. According to our current understanding, the ideal platelet concentration for tissue repair is 5–10× baseline, 1.5–3 million/μL.
Stem Cells
Stem cells are unspecialized cells distinguished by two unique characteristics: 1) they are capable of self-renewal through division and 2) under certain physiologic or experimental conditions, they can be induced to become tissue- or organ-specific cells. Most organ systems of the body have a resident pool of somatic, tissue-specific stem cells, in many cases of traumatic injury or disease, the quantity and potency of endogenous stem cell populations are insufficient to regenerate compromised tissues. ,
Embryonic stem cells (ESCs) are stem cells derived from the undifferentiated inner mass cells of a human embryo. ESCs are pluripotent, meaning they are able to grow (i.e., differentiate) into all derivatives of the three primary germ layers: ectoderm, endoderm, and mesoderm. These are currently only used for research purposes.
MSCs have a narrower range of potential and can differentiate into a variety of cell types, including osteoblasts, chondrocytes, myocytes, and adipocytes. MSCs can be derived from a variety of sources, though the primary sites of extraction have been bone marrow and adipose tissue. Stem cells, in general are relatively scarce; they can be difficult to isolate; grow slowly; and do not differentiate well without appropriate peripheral cytokines. It can be further difficult to isolate sufficient amounts required for therapy, which is worse depending on the source.
The therapeutic activity of MSCs is mediated by paracrine effects. Whereas the common misconception is that the stem cells will identify injured tissues and differentiate directly into those tissues by engraftment, it has been demonstrated that less than 1% of stem cells injected remain after as little as 7 days. MSCs respond to injured tissues and can coordinate a healing response via secretion of bioactive molecules, such as Ang-1, Ang-2, BMP, BDNF, IL-6, and VEGF.
Osteoarthritis
A growing body of evidence has accumulated examining PRP and prolotherapy as a treatment of knee OA. Several studies now have demonstrated that intraarticular PRP injections are a safe and effective treatment to reduce pain and improve quality of life through increased function. The autologous nature of PRP and autologous stem cells theoretically reduces the risk of potential side effect associated with alternative injectates such as corticosteroids and hyaluronic acid (HA). A systematic review of all the studies comparing PRP to HA identified 12 studies, 10 of which were prospective randomized controlled studies, which found PRP to be superior to HA in reduction of pain, as well as demonstrating an improvement in functional outcome measures.
Prolotherapy targets include knee stabilizing ligaments and intraarticular injections. Systematic review of 3 high quality RCTs, a total of 258 patients proved prolotherapy beneficial at 24 weeks, with one study following the patients up to a year and confirmed similarly better WOMAC and VAS scores.
Dextrose prolotherapy has been also shown to benefit temporomandibular joint hypermobility and associated pain.
Tendinopathy
Tendinopathies are the various conditions associated with tendon pain primarily caused by overuse. Tendinopathy is associated with histopathologic changes such as minimal inflammation, degeneration and disorganization of collagen fibers, and increased cellularity. Macroscopic changes include pain, tendon thickening, and the loss of structural integrity. Tendon overuse leads to an imbalance between the protective/regenerative changes of the tissue and pathologic responses from overuse, which results in pain, tearing, weakness, and degeneration. Tendons act as an interface between muscles and the skeletal structures. When tendons are exposed to supramaximal loading, injury occurs. A tendons’ intrinsic low metabolic rate may also lead to delayed wound healing when injury does occur.
With age, changes in collagen structure, such as a loss of water content, predispose tendons to damage. Vascularity also decreases with age, and tendon disease often occurs at these hypovascular areas. Instability or impingement leads to abnormal and excessive loading of the tendon which predisposes to injury. Collagen fibrils can rupture and these regions may together form intrasubstance tears. These intrasubstance tears may extend to the surface, eventually progressing to full thickness tears. Generally, degenerative changes occur before macroscopic tendon tears develop and as such, it is unusual for a tear to occur in a nondegenerated tendon.
Level 1 evidence supports the use of PRP in the treatment of lateral epicondylopathy of the elbow (Tennis elbow) with superiority in pain relief and functional outcomes to corticosteroids up to 24-month follow-up. Evidence also supports the use of RM in the treatment of Achilles, patellar, and hamstring tendinopathy.
Ligament Pathology
Ligaments are dense connective tissue that connect bone to bone and provide stabilization to a joint. Though ligaments are functionally different from tendons as they connect bone to bone, they are structurally similar. The main differences are that ligaments have higher proteoglycan content, higher water content, lower in collagen content, and are less uniform. These structures are typically injured with supraphysiologic stretching, at the end range of motion for a joint. Acute trauma typically causes ligament abnormalities and is often marked by fluid surrounding the ligament, although chronic repetitive microtrauma may be a factor as with tendon injuries. Potential damage includes interstitial tearing of collagen fibers, partial tears that extend to the surface, and full thickness ligament ruptures. Over time, the ligament can become elongated and lax. Other evidence of injuries includes bone contusions, fractures, or joint effusion. After healing, the ligament may appear thickened, weakened, and prone to further damage. Common examples include the anterior talofibular ligament in ankle sprains and anterior cruciate ligament in knee injuries. There are limited studies investigating RM for ligamentous injuries, but there is evidence that PRP may promote the success rate of ACL repairs and provide pain relief in plantar fasciitis superior to corticosteroid. , Prolotherapy has been successfully used for sacroiliac joint related pain, low back pain, and coccydynia.
Adjunct Medications
Local anesthetics, corticosteroids, and contrast agents are routinely used during interventional orthopedic and pain management procedures for both diagnostic and therapeutic purposes. A growing body of literature suggests that these routinely used injectates promote catabolic processes including apoptosis which are thought to accelerate the disease process. It is therefore paramount to understand the effect of these agents on RM injectates, and on target tissues including tenocytes, chondrocytes, nucleus pulposus, and ligamentous tissue. Numerous studies have shown time- and dose-dependent chondrotoxicity of local anesthetics on human and animal soft tissues, with ropivacaine likely being the least toxic offender.
Contrast agents are considered necessary for some procedures to confirm safe and accurate needle placement. The use of contrast agents can be avoided by basing accurate needle placement on radiographic imaging or utilizing alternative image guidance such as ultrasound. Contrast agents exerted chondrotoxic effects in a dose- and type-dependent manner with ionic contrasts being most detrimental. Nonionic demonstrated mild dose-dependent chondrotoxicity and were the least harmful of those studied.
Conclusion
RM is an emerging field of medicine that demonstrates Level 1 evidence in the treatment of common musculoskeletal diseases. Further research is necessary to elucidate the parameters that optimize outcomes, including patient selection, pre- and postinjection protocols, and procurement of RM injectate.
High Yield Points
RM encompasses an emerging field medicine with the goal of replacing, engineering, or regenerating human cells, tissues, or organs lost or injured due to age, disease, or congenital defects to restore or establish normal function.
Dextrose prolotherapy creates tissue injury and with that initiates the healing cascade, PRP itself is the start of the healing cascade with the degranulation of the platelets. This inflammatory response by prolotherapy or PRP also attracts MSCs to further synchronize healing, while there is also an option to directly inject MSCs with PRP acquired from bone marrow concentrate. Fig. 17.1 .
