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Joint articular cartilage has very little capacity for self-regeneration. Areas of articular cartilage damage tend to get worse over time rather than better. This process often leads to the spiral of joint destruction ending in arthritis. The mature cells within the cartilage (chondrocytes) lack the capacity to produce cartilage matrix to effectively repair defects. The quest for cells that contain the ability to produce reparative matrix has led scientists to mesenchymal STEM cells (MSCs) as a potential source.
A stem cell is a progenitor cell the can transform (or “differentiate”) into various cell types, depending on the environment in which it is placed. What makes stem cells an appealing possibility for cartilage repair is their ability to transform into chondrocytes when placed in the correct cartilage environment?
Stem cells can be found in many areas of the body (muscle, blood, adipose), though a common source is from bone marrow. Bone marrow is accessed through the rim of the pelvis known as the iliac crest. Bone marrow stems cells are the mesenchymal stem cells, or MSCs. MSCs are aspirated with a needle under local anesthesia, concentrated, and then applied to the cartilage defect in the knee. It may be preferable that these cells are expanded into higher quantities in the laboratory for several weeks (increasing their generation time) before they are inserted into the joint.
MSCs should be distinguished from other types of stem cells; embryonic stem cells, or ESCs. MSCs do not carry the ethical concerns of ESCs.
Wharton’s Jelly MSCs, derived form the umbilical cord of a new born.
Hematopoetic stems cell, which are derived from the peripheral blood.
The exact mechanism of stem cell based regeneration is not completely elucidated. Whether MSCs differentiate directly into cartilage cells, or do they somehow “regulate” cartilage behavior is up for debate. We think that stem cells “talk to“ chondrocytes.
A high level of oxygen tension is detrimental to stem cells. These cells prefer environments of lower levels of oxygen.
The age of the patient may play a role in the potency of the stem cells. We know that stem cell vigor decreases with age.
There are many variables that may impact the outcome of stem cell treatment, most importantly, the type and severity of cartilage lesion, i.e., acute vs. chronic, location and depth.
One concern is that stem cells favor differentiation towards bone rather than cartilage, sometimes causing unwanted bone formation. This process may require modulation of the stem cell mechanism with various growth mediators.
The development of scaffolds to contain the stem cells in the cartilage defect is an area of much investigation. The simple injection of stem cells in a damaged joint is unlikely to confer a great benefit. The goal is to apply the cells to the specific area of cartilage injury. There are various mediums being investigated to do this.
“There are now several reports of adult stem cells in other tissues (muscle, blood, and fat) that demonstrate plasticity. Very few published research reports on plasticity of adult stem cells have, however, included clonality studies. That is, there is limited evidence that a single adult stem cell or genetically identical line of adult stem cells demonstrates plasticity” (Adult Stem Cell).
Stem cells are usually collected by aspirating bone marrow from the iliac crest, which is the edge of the pelvis above the hip. The bone marrow aspirate is a bloody substance that is removed from the pelvis through a small needle. The area is well anesthetized with a local anesthetic and there is surprisingly little discomfort with this procedure. Most procedures require about 60cc of bone marrow aspirate (one large syringe). This aspirate contains mesenchymal stem cells, platelets, and other types of stem cells.
The bone marrow is placed in a centrifuge. The centrifuge spins the bone marrow at a very fast speed causing the stem cells and platelets to be separated from the rest of the blood products. It is this concentration of bone marrow that is filtered and injected back into the joint or injured area. The concentration is called BMAC or bone marrow aspiration concentrate.
NIH website (http://stemcells.nih.gov/info/basics/basics4.asp) has a thorough explanation, as follows:
What are adult stem cells?
An adult stem cell is thought to be an undifferentiated cell, found among differentiated cells in a tissue or organ that can renew itself and can differentiate to yield some or all of the major specialized cell types of the tissue or organ. The primary roles of adult stem cells in a living organism are to maintain and repair the tissue in which they are found. Scientists also use the term somatic stem cell instead of adult stem cell, where somatic refers to cells of the body (not the germ cells, sperm or eggs). Unlike embryonic stem cells, which are defined by their origin (cells from the preimplantation-stage embryo), the origin of adult stem cells in some mature tissues is still under investigation.
Research on adult stem cells has generated a great deal of excitement. Scientists have found adult stem cells in many more tissues than they once thought possible. This finding has led researchers and clinicians to ask whether adult stem cells could be used for transplants. In fact, adult hematopoietic, or blood-forming, stem cells from bone marrow have been used in transplants for 40 years. Scientists now have evidence that stem cells exist in the brain and the heart. If the differentiation of adult stem cells can be controlled in the laboratory, these cells may become the basis of transplantation-based therapies.
The history of research on adult stem cells began about 50 years ago. In the 1950s, researchers discovered that the bone marrow contains at least two kinds of stem cells. One population, called hematopoietic stem cells, forms all the types of blood cells in the body. A second population, called bone marrow stromal stem cells (also called mesenchymal stem cells, or skeletal stem cells by some), were discovered a few years later. These non-hematopoietic stem cells make up a small proportion of the stromal cell population in the bone marrow, and can generate bone, cartilage, fat, cells that support the formation of blood, and fibrous connective tissue.
In the 1960s, scientists who were studying rats discovered two regions of the brain that contained dividing cells that ultimately become nerve cells. Despite these reports, most scientists believed that the adult brain could not generate new nerve cells. It was not until the 1990s that scientists agreed that the adult brain does contain stem cells that are able to generate the brain's three major cell types—astrocytes and oligodendrocytes, which are non-neuronal cells, and neurons, or nerve cells.
A. Where are adult stem cells found, and what do they normally do?
Adult stem cells have been identified in many organs and tissues, including brain, bone marrow, peripheral blood, blood vessels, skeletal muscle, skin, teeth, heart, gut, liver, ovarian epithelium, and testis. They are thought to reside in a specific area of each tissue (called a "stem cell niche"). In many tissues, current evidence suggests that some types of stem cells are pericytes, cells that compose the outermost layer of small blood vessels. Stem cells may remain quiescent (non-dividing) for long periods of time until they are activated by a normal need for more cells to maintain tissues, or by disease or tissue injury.
Typically, there is a very small number of stem cells in each tissue, and once removed from the body, their capacity to divide is limited, making generation of large quantities of stem cells difficult. Scientists in many laboratories are trying to find better ways to grow large quantities of adult stem cells in cell culture and to manipulate them to generate specific cell types so they can be used to treat injury or disease. Some examples of potential treatments include regenerating bone using cells derived from bone marrow stroma, developing insulin-producing cells for type 1 diabetes, and repairing damaged heart muscle following a heart attack with cardiac muscle cells.
B. What tests are used for identifying adult stem cells?
Scientists often use one or more of the following methods to identify adult stem cells:
(1) label the cells in a living tissue with molecular markers and then determine the specialized cell types they generate; (2) remove the cells from a living animal, label them in cell culture, and transplant them back into another animal to determine whether the cells replace (or "repopulate") their tissue of origin.
Importantly, it must be demonstrated that a single adult stem cell can generate a line of genetically identical cells that then gives rise to all the appropriate differentiated cell types of the tissue. To confirm experimentally that a putative adult stem cell is indeed a stem cell, scientists tend to show either that the cell can give rise to these genetically identical cells in culture, and/or that a purified population of these candidate stem cells can repopulate or reform the tissue after transplant into an animal.
C. What is known about adult stem cell differentiation?
As indicated above, scientists have reported that adult stem cells occur in many tissues and that they enter normal differentiation pathways to form the specialized cell types of the tissue in which they reside.
Normal differentiation pathways of adult stem cells. In a living animal, adult stem cells are available to divide, when needed, and can give rise to mature cell types that have characteristic shapes and specialized structures and functions of a particular tissue. The following are examples of differentiation pathways of adult stem cells (Figure 2) that have been demonstrated in vitro or in vivo.
Transdifferentiation. A number of experiments have reported that certain adult stem cell types can differentiate into cell types seen in organs or tissues other than those expected from the cells' predicted lineage (i.e., brain stem cells that differentiate into blood cells or blood-forming cells that differentiate into cardiac muscle cells, and so forth). This reported phenomenon is called transdifferentiation.
Although isolated instances of transdifferentiation have been observed in some vertebrate species, whether this phenomenon actually occurs in humans is under debate by the scientific community. Instead of transdifferentiation, the observed instances may involve fusion of a donor cell with a recipient cell. Another possibility is that transplanted stem cells are secreting factors that encourage the recipient's own stem cells to begin the repair process. Even when transdifferentiation has been detected, only a very small percentage of cells undergo the process.
In a variation of transdifferentiation experiments, scientists have recently demonstrated that certain adult cell types can be "reprogrammed" into other cell types in vivo using a well-controlled process of genetic modification (see Section VI for a discussion of the principles of reprogramming). This strategy may offer a way to reprogram available cells into other cell types that have been lost or damaged due to disease. For example, one recent experiment shows how pancreatic beta cells, the insulin-producing cells that are lost or damaged in diabetes, could possibly be created by reprogramming other pancreatic cells. By "re-starting" expression of three critical beta-cell genes in differentiated adult pancreatic exocrine cells, researchers were able to create beta cell-like cells that can secrete insulin. The reprogrammed cells were similar to beta cells in appearance, size, and shape; expressed genes characteristic of beta cells; and were able to partially restore blood sugar regulation in mice whose own beta cells had been chemically destroyed. While not transdifferentiation by definition, this method for reprogramming adult cells may be used as a model for directly reprogramming other adult cell types.
In addition to reprogramming cells to become a specific cell type, it is now possible to reprogram adult somatic cells to become like embryonic stem cells (induced pluripotent stem cells, iPSCs) through the introduction of embryonic genes. Thus, a source of cells can be generated that are specific to the donor, thereby avoiding issues of histocompatibility, if such cells were to be used for tissue regeneration. However, like embryonic stem cells, determination of the methods by which iPSCs can be completely and reproducibly committed to appropriate cell lineages is still under investigation.
D. What are the key questions about adult stem cells?
Many important questions about adult stem cells remain to be answered. They include:
- How many kinds of adult stem cells exist, and in which tissues do they exist?
- How do adult stem cells evolve during development and how are they maintained in the adult? Are they "leftover" embryonic stem cells, or do they arise in some other way?
- Why do stem cells remain in an undifferentiated state when all the cells around them have differentiated? What are the characteristics of their “niche” that controls their behavior?
- Do adult stem cells have the capacity to transdifferentiate, and is it possible to control this process to improve its reliability and efficiency?
- If the beneficial effect of adult stem cell transplantation is a trophic effect, what are the mechanisms? Is donor cell-recipient cell contact required, secretion of factors by the donor cell, or both?
- What are the factors that control adult stem cell proliferation and differentiation?
- What are the factors that stimulate stem cells to relocate to sites of injury or damage, and how can this process be enhanced for better healing?
Results in the Literature
The objective, which is yet to be achieved by any method, is natural tissue repair. MSCs are known to secrete bioactive molecules that stimulate angiogenisis and mitosis of intrinsic progenitor cells. The concept is that once the stem cells are introduced back into the body, they will take signals from their environment and nourishment from any remaining vasculature or blood supply and start to grow, differentiating into bone or cartilage. This new tissue regenerates into the scaffold of the surrounding tissue. Though the regenerated tissue may lack the material characteristics of normal tissue, it is hoped that the joint longevity can be prolonged, improving symptoms of pain and postponing further procedures such as replacement. A long-standing concern is phenotypic stability of MSCs, that is, that tendency of the stem cells to transform into the desired tissue like cartilage and not an undesirable tissue like bone.
To date, no study has demonstrated that normal, organized cartilage can be completely regenerated. However recent studies have elucidated the promise of stem cells in musculoskeletal tissue:
Ochi et al (2004) observed in a rat model that the injection of cultured MSCs combined with microfracture could accelerate the regeneration of cartilage and concluded that this approach could represent an effective and less invasive strategy for the regeneration of articular surfaces.
Giannini et al (2009, CORR) used MSCs to address osteochondral lesion of the ankle talus. The technique was used in 48 patients with 2 year follow-up. Almost a 50% improvement was seen in pain and function scores. Non-hyaline cartilage formation was the rule.
Niemeyer et al (2010, Biomaterials) used a sheep model to study the effects of MSCs on bone healing. A significantly higher healing capacity was seen in those animals utilizing BMAC derived stem cells. Adding PRP to the composite further enhance healing effects.
Hernigou (JBJS) in a study evaluating percutaneus injection of stem cells to enhance bone healing concluded that percutaneous autologous bone-marrow grafting is an effective and safe method for the treatment of anatrophic tibial diaphyseal nonunion. However, its efficacy appears to be related to the number of progenitors in the graft, and the number of progenitors available in bone marrow aspirated from the iliac crest appears to be less than optimal in the absence of concentration.
In one of the more significant stem cell studies, Fortier et al (2010, JBJS) studied the effects of BMAC injections in chondral lesions using a horse model. Full thickness cartilage lesions were created in 15 horse knees. The defects were treated with BMAC and microfracture or microfrature alone. At 8 months follow-up, MRI and histological scores demonstrated increased fill of the defects and integration of repair tissue into surrounding cartilage in the BMAC group. There was greater type II hyaline cartilage content in the BMAC group as well. They concluded that using BMAC along with microfracture can result in healing of acute full-thickness cartilage defects that is superior to that after microfracture alone in an equine model.
In a very promising study, Gobbi et al (2011, Cartilage) reported on 15 surgical patients with large chondral injuries in the knee (av 9.2mm2), which were repaired with a combination of BMAC and collagen matrix through an open incision. Significant improvement in pain and function scores was seen at 2 years of follow-up. Smaller lesion faired better. Biopsies done in 3 patients showed a mixture of hyaline and fibrocartilage growth, with favorable organization characteristics (preferable to ACI). There was no control group in this study and the authors concede that some level of improvement would likely have been seen with procedures such as microfracture, yet this remains a convincing report on the potential of stem cells.