The Ideal Recipe for Cartilage from Stem Cells


Researchers at Case Western Reserve and Harvard University will use a 5-year, $2-million NIH grant to build a microfactory that bangs out the optimal formula for joint cartilage. Such an end product could one day potentially benefit many of the tens of thousands of people in the United States who suffer from cartilage loss or damage.

Joint cartilage or articular cartilage caps the ends of long bones and bears the loads, absorbs shocks and, in combination with lubricating synovial fluid, helps knees, hips, shoulders, and other joints to smoothly bend, lift, and rotate. Unfortunately, this tissue has little capacity to regenerate, which means that there is a critical need for new therapeutic strategies.

Artificial substitutes cannot match real cartilage and attempts to engineer articular cartilage have been stymied by the complexities of directing stem cells to differentiate into chondrocytes and form the right kind of cartilage.

Stem cells are quite responsive to the environmental cues presented to them from their surroundings. What this research project hopes to determine are those specific cue that drive stem cells to differentiate into chondrocytes that make the right kind of cartilage with the right kind of microarchitecture that resembles natural, articular cartilage. To do this, they will engage in a systematic study of the effects of cellular micro-environmental factors that influence stem cell differentiation and cartilage formation.

Bone marrow- and fat-derived mesenchymal stem cells have been differentiated into cartilage-making chondrocytes in the laboratory. These two stem cell populations are distinct, however, and required different conditions in order to drive them to differentiate into chondrocytes. This research group, however, has designed new materials with unique physical properties, cell adhesive capabilities, and have the capacity to deliver bioactive molecules.

By controlling the presentation of these signals to cells, independently and in combination with mechanical cues, this group hopes to identify those most important cues for driving cells to differentiate into chondrocytes.

Ali Khademhosseini specializes in microfabrication and micro-and nano-scale technologies to control cell behavior. He and his team will develop a microscale high-throughput system at his laboratory that will accelerate the testing and analysis of materials engineered in another laboratory.

This research cooperative hopes to test and analyze more than 3,000 combinations of factors that may influence cell development, including differentiation, amounts of biochemicals, extracellular matrix properties, compressive stresses, and more. Khademhosseini and his colleagues hope to begin testing comditions identified from these studies in animal models by the of the grant term.

Non-Randomized Stem Cell Study for Knee Osteoarthritis Yields Positive Results


A peer-reviewed study that was neither placebo-controlled nor randomized, but did examine 840 patients, has shown that the use of a patient’s own bone marrow stem cells are both safe and effective.

Christopher Centeno and his colleagues, who pioneered the Regenexx protocol, use live-imaging to guide the application of stem cells to the site in need of healing. Centeno and others have established several clinics around the United States that utilize the Regenexx system, and the data published in this paper came from these clinics, in addition to Chris Centeno’s own clinic in the Denver, Colorado area.

In this study, patients self-rated their lower extremity functional using a lower extremity functional scale (LEFS), and their knee pain by using a numerical pain scale (NPS). Patients had bone marrow extracted through a bone marrow aspiration. These bone marrow cells were isolated and concentrated, and then prepared for reinvention. In addition, platelet rich plasma (PRP) and platelet lysate (PL) were prepared from the patient’s own blood and these, with the bone marrow cells, were injected into the knee under guided imaging. The frequency and types of adverse events (AE) were also recorded by the physicians.

Some of these patients had fat overlaid on their knee lesions in addition to their bone marrow cells. Of the 840 procedures that were performed, 616 had treatment without additional fat, and 224 had treatment with the fat graft. This was to determine if the use of fat, with its resident stem cell population, augmented healing of the arthritic knee.

When the LEFS scores before and after the Regenexx procedure were compared, an increase of 7.9 and 9.8 in the two groups (out of 80) was observed. The mean NPS score decreased from 4 to 2.6 and from 4.3 to 3 in the two groups. AE rates were 6% and 8.9% in the two groups. An examination of these data showed that pre- and posttreatment improvements were statistically significant. However, the differences between the fat- and fat+ groups were statistically insignificant.

The patients in this study suffered from osteoarthritis. Consequently, they were experiencing significant knee pain and many were candidates for a knee replacement. Many of these patients were able to avoid knee replacement by undergoing the Regenexx procedure.

The study concluded that there was no advantage of adding fat to the joint over the bone marrow cells. Safety in both groups (with and without fat) was excellent compared to knee replacement.

This study used data from patients who were part of the Regenexx registry. Therefore, this study was not a randomized, controlled study, like the kind that are used to test drugs. Randomized controlled trials are being conducted by Centeno and his colleagues at the various Regenexx centers. A knee osteoarthritis study is being studied in Chicago, another study regarding shoulder rotator cuff tears, and a third study examining ACL tears are in progress.

Cartilage Repair Using Human Umbilical Cord Blood-Derived Mesenchymal Stem Cells Embedded in Hyaluronic Acid Hydrogel in a Minipig Model


Cartilage shows lousy regenerative capabilities. Fortunately, it is possible to regenerate cartilage with human umbilical cord blood-derived mesenchymal stem cells (hUCB-MSCs) that have been embedded in a hyaluronic acid (HA) hydrogel composite. In fact, such a combination has shown remarkable results in rat and rabbit models.

In this present study, published in Stem Cells Translational Medicine, Yong-Geun Park and his colleagues from SungKyunKwan University School of Medicine, in Seoul, South Korea sought to confirm the efficacy of this protocol in a in a pig model using three different hUCB-MSC cell lines.

Park and his coworkers generated full-thickness cartilage injuries in the trochlear groove of each knee in 6 minipigs. Three weeks later, an even larger cartilage defect, 5 mm wide by 10 mm deep, was created, followed by an 8-mm-wide and 5-mm-deep boring. In short, the knee cartilages of these minipigs were very messed up.

Trochlear-groove

To these knee cartilages, a mixture (1.5 ml) of hUCB-MSCs (0.5 × 107 cells per milliliter) and 4% HA hydrogel composite were troweled into was then cartilage defects of the right knee. The left knee served as an untreated control. Each cell line was used in two minipigs.

Macroscopic findings of the osteochondral defects of the porcine knees. At 12 weeks postoperatively, the defects of both knees had produced regenerated tissues that were pearly white and firm. These new tissues, which resembled articular cartilage, appeared adherent to the adjacent cartilage and had restored the contour of the femoral condyles (smooth articular surfacewithout depression). The regenerated tissue of the control knee (left knee) looked fibrillated. Grossly, no differencewas seen in the quality of the repaired tissue in the transplanted knee (right knee) among the three groups with different cell lines. (A): Group A. (B): Group B. (C): Group C. Abbreviations: HA, hyaluronic acid; hUCB-MSCs, human umbilical cord blood-derived mesenchymal stem cells.
Macroscopic findings of the osteochondral defects of the porcine knees. At 12 weeks postoperatively, the defects of both
knees had produced regenerated tissues that were pearly white and firm. These new tissues, which resembled articular cartilage, appeared adherent to the adjacent cartilage and had restored the contour of the femoral condyles (smooth articular surface without depression). The regenerated tissue of the control knee (left knee) looked fibrillated. Grossly, no difference was seen in the quality of the repaired tissue in the transplanted knee (right knee) among the three groups with different cell lines. (A): Group A. (B): Group B. (C): Group C. Abbreviations: HA, hyaluronic acid; hUCB-MSCs, human umbilical cord blood-derived mesenchymal stem cells.

12 weeks after surgery, the pigs were sacrificed, and the degree of subsequent cartilage regeneration was evaluated by gross and more detailed microscopic analysis of the knee tissue. The transplanted knee showed superior and more complete joint-specific (hyaline) cartilage regeneration compared with the control knee. The microscopic characteristics of the knee cartilage showed that those animals that received the hUCB-MSCs had greater rates of cell proliferation and cells that differentiated into cartilage-making cells.

Microscopic findings of the regenerating osteochondral defects on porcine articular cartilage (safranin O and fast green staining). At 12 weeks postoperatively, the surface of the repairing tissue in the control knee (left knee) was poorly stained for glycosaminoglycan. In the transplanted knee (right knee), both the regenerated tissue and the adjacent cartilage to which it had become adherent exhibited the normal orthochromatic staining properties with safranin O. (A): Group A. (B): Group B. (C): Group C. Scale bars = 2 mm. Abbreviations: HA, hyaluronic acid; hUCB-MSCs, human umbilical cord blood-derived mesenchymal stem cells.
Microscopic findings of the regenerating osteochondral defects on porcine articular cartilage (safranin O and fast green staining). At 12 weeks postoperatively, the surface of the repairing tissue in the control knee (left knee) was poorly stained for glycosaminoglycan. In the transplanted knee (right knee), both the regenerated tissue and the adjacent cartilage to which it had become adherent exhibited the normal orthochromatic staining properties with safranin O. (A): Group A. (B): Group B. (C): Group C. Scale bars = 2 mm. Abbreviations: HA, hyaluronic acid; hUCB-MSCs, human umbilical cord blood-derived mesenchymal stem cells.

These data show consistent cartilage regeneration using composites of hUCB-MSCs and HA hydrogel in a large animal model. These experiments could be a stepping stone to a human clinical trial in the future that treats osteoarthritis of the knees with hUCB-MSCs embedded in HA hydrogel.

Cartilage-Making Stem Cells from Joints


Chiharo Akazawa from the Tokyo Medical and Dental University and his colleagues have tested two types of mesenchymal stem cells from human patients for their ability to make bone, cartilage, or fat. Their tests illustrated what has been shown several time before; mesenchymal stem cells tend to differentiate into the tissues that most closely resemble their tissue of origin.

Akazawa and his colleagues previously discovered a way to effectively isolated mesenchymal stem cells from bone marrow, which is no small feat because mesenchymal stem cells (MSCs) are a minority of the cells in bone marrow (Mabuchi and others (2013), Stem Cell Reports 1: 152-165). In a recent paper in the journal PLoS ONE, Akazawa and others used this technique to isolate MSCs from bone marrow and from synovial membrane – the fluid-filled sac that encases joints. In large joints, this synovium is large and called a “bursa.”.

In culture, the bone marrow-derived MSCs from several different human donors showed a marked tendency to form bone, but they did not make good cartilage or fat. The synovial MSCs, on the other hand, did not do so well at making bone, but made very good fat and cartilage. These differentiation trends were observed in MSCs culture for several different human donors. All cells were collected during arthroscopic surgery.

Since the synovial membrane of patients suffering from osteoarthritis undergoes, increased cell division, it is possible that the number of stem cells also increases. Alternatively, using MSCs from healthy donors who do not have arthritis may be even more preferable. Nevertheless, MSCs from synovial membrane show excellent cartilage-making potential and they may be a suitable source of cell for cartilage regeneration.

Mesoblast Phase Degenerative Disc Disease Treatment Receives Positive Feedback from European Regulatory Agencies


Mesoblast Limited announced that European Medicines Agency has approved expansion of their Phase 3 clinical program of its product candidate MPC-06-1D for degenerative disc disease.

Mesoblast’s Phase 3 program for this product candidate is currently in the process of enrolling patients in the United States under an Investigational New Drug (IND) application filed with the US Food and Drug Administration (FDA).  Having received general agreement from EMA on the target patient population, trial size, primary composite endpoint, and comparators in the control population, Mesoblast now intends to additionally enroll patients across multiple European sites.

The discussions with EMA occurred as part of combined scientific and reimbursement advice under an EU pilot program known as Shaping European Early Dialogues (SEED). The SEED pilot program was established to facilitate early dialogue between EMA, European Health Technology Assessment reimbursement bodies, and selected companies with late-stage clinical development programs. Mesoblast’s product candidate MPC-06-ID is one of only seven medicines accepted for the SEED program.

Mesoblast and SEED representatives discussed key clinical trial aspects of the development of MPC-06-ID including the safety database, mechanisms of action, patient population and trial size, composite endpoints, and comparators. The discussions also focused on access to EU markets and pharmacoeconomic endpoints that may lead to reimbursement.

The guidance from the meeting with SEED representatives may result in a final comprehensive EU development and commercialization program that has an increased likelihood of producing data that will be acceptable for both registration and reimbursement review in multiple European countries.

High-Quality Cartilage Production from Pluripotent Stem Cells


High-quality cartilage has been produced from pluripotent stem cells by workers in the laboratory of Sue Kimber and her team in the Faculty of Life Sciences at The University of Manchester. Such success might be used in the future to treat the painful joint condition osteoarthritis.

Kimber and her colleagues used strict laboratory conditions to grow and transform embryonic stem cells into cartilage cells known as chondrocytes.

Professor Kimber said: “This work represents an important step forward in treating cartilage damage by using embryonic stem cells to form new tissue, although it’s still in its early experimental stages.” Kimber’s research was published in Stem Cells Translational Medicine.

During the study, the team analyzed the ability of embryonic stems cells to become cartilage precursor cells. Kimber and her colleagues then implanted these pre-chrondrocytes into cartilage defects in the knee joints of rats. After four weeks, the damaged cartilage was partially repaired and following 12 weeks a smooth surface, which looked very similar to normal cartilage, was observed. More detailed studied of this newly regenerated cartilage demonstrated that cartilage cells from embryonic stem cells were still present and active within the tissue.

Developing and testing this protocol in rats is the first step in generating the information required to run such a study in people with arthritis. Before such a clinical trial can be run, more data will need to be collected in order to check that this protocol is effective and that there are no toxic side-effects.

However, Kimber and her coworkers say that this study is very promising as not only did this protocol generate new, healthy-looking cartilage but also importantly there were no signs of any side-effects such as growing abnormal or disorganized, joint tissue or tumors. Further work will build on this finding and demonstrate that this could be a safe and effective treatment for people with joint damage.

Chondrocytes created from adult stem cells are being used on an experimental basis, but, to date, they cannot be produced in large amounts, and the procedure is expensive.

With their huge capacity to proliferate, pluripotent stem cells such as embryonic stem cells and induced pluripotent stem cells can be manipulated to form almost any type of mature cell. Such cells offer the possibility of high-volume production of cartilage cells, and their use would also be cheaper and applicable to a greater number of arthritis patients, the researchers claim.

“We’ve shown that the protocol we’ve developed has strong potential for developing large numbers of chondrogenic cells appropriate for clinical use,” added Prof Kimber. “These results thus mark an important step forward in supporting further development toward clinical translation.”

Osteoarthritis affects more than eight million people in the UK alone, and is a major cause of disability. It and occurs when cartilage at the ends of bones wears away causing joint pain and stiffness.

Director of research at Arthritis Research UK Dr Stephen Simpson added: “Current treatments of osteoarthritis are restricted to relieving painful symptoms, with no effective therapies to delay or reverse cartilage degeneration. Joint replacements are successful in older patients but not young people, or athletes who’ve suffered a sports injury.

“Embryonic stem cells offer an alternative source of cartilage cells to adult stem cells, and we’re excited about the immense potential of Professor Kimber’s work and the impact it could have for people with osteoarthritis.”

European Knee Meniscus Injury Pilot Trial to Evaluate Cytori Cell Therapy Begins


Cytori Therapeutics is a cell therapy company that is in the process of developing cell therapies from a patient’s own fat tissue that can potentially treat a variety of medical conditions. To date, the preclinical studies and clinical trials suggest that their Cytori Cell Therapy can improve blood flow, modulate the immune system, and facilitating wound repair.

Recently, Cytori has announced that it has enrolled its first patients in an ambitious clinical trial that will test their stem cell product in patients undergoing surgery for traumatic injuries to the meniscus of the knee.  The meniscus is a wedge of cartilage on either side of the knee joint that acts a a shock absorber between the femur and the head of the tibia.

meniscus

Ramon Cugat, who is the Co-Director of the Orthopedic Institute, Hospital Quiron Barcelona, Spain, is the principal investigator for this trial. Dr. Cugat serves as an orthopedic surgeon at Hospital Quiron Barcelona. This trial will test the ability of Cytori Cell Therapy to heal the meniscus and is being conducted in parallel with a similar trial that is testing the Cytori Cell Therapy as a treatment for anterior cruciate ligament (ACL) repairs. The patients treated with Cytori Cell Therapy for ACL repairs are still being evaluated, but to date, no safety related concerns have emerged and the patients seem to have improved. These preliminary results were presented at the Barcelona Knee Symposium in November 2014.

knee_joint

“Dr. Cugat is a leading expert in treating traumatic knee injuries in elite athletes,” said Dr. Marc H. Hedrick, President and CEO of Cytori Therapeutics. “These trials are important to Cytori because, at minimal cost to us, they provide additional clinical evidence that our therapy can be safely used in treating a multitude of knee conditions.”

The meniscus trial is a two-center, phase I study that will assess the safety and efficacy of Cytori’s ECCM-50 adipose-derived regenerative cell therapy in meniscus repair. In this trial, up to 60 patients who have had meniscus surgery to repair the meniscus will receive injection of the cells directly into the meniscus. Each patient will be evaluated by several clinical read outs that assess the recovery of the patient after meniscus surgery. As in the case of the ACL repair study, the goal of this trial is to determine if Cytori Cell Therapy can be safely delivered to the meniscus and whether efficacy can be observed.

“Tears to the meniscus are problematic injuries for active individuals, particularly athletes. Based on the early results from a recent series of 20 patients treated for complete anterior cruciate ligament injuries, we are eager to evaluate whether augmentation surgery with Cytori Cell Therapy will lead to quicker and more complete healing,” said Dr. Cugat.

Injected patients will fill out a patient questionnaire that assesses knee pain, function and activity, This questionnaire is called the Knee Injury and Osteoarthritis Outcome Score (KOOS), but patients will also be physically examined to ascertain the extent of their knee function and the degree of their movement, with or without pain. Patients will be given a visual analogue score to assess knee pain, and knee function will be assessed by the Lysholm Knee Scoring Scale, Tegner Activity Scale, and the Lower Extremity Functional Scale. Each patient will also have their knees examined by Magnetic Resonance Imaging (MRI) in order to examine the structural integrity of their meniscus. These assessments will be taken before and 60, 90, 180 and 365 days after surgery and the MRIs will be done before and at 90, 180 and 365 days after surgery.

The preliminary results of the ACL study showed that the Cytori strategy was feasible and did not result in any significant safety issues above that seen with a standard small volume liposuction. All the injected patients recovered without any complications. The results of the ACL trial were compared to a historical control group of patients who had the same surgical procedure by the same surgical team but without other interventions. Overall, the patient’s recovery from pain and their ability to return to daily activities was accelerated as a result of the therapeutic enriched bone-patellar tendon-bone transplant. Both the patient questionnaires and serial MRI scans of the knees following cell therapy were consistent with accelerated healing of the graft. Presently, Dr. Cugat and his coworkers are obtaining one year follow-up information on the treated patients and they will report their data in a peer-reviewed journal in the future.

ACL and meniscus tears are among the most common sports-related knee injuries and unfortunately, these two injuries often are sustained simultaneously. According to the American Academy of Orthopedic Surgeons, ACL injuries have an annual incidence of more than 200,000 cases with nearly half undergoing surgical reconstruction. Further, an estimated 850,000 patients undergo surgical procedures to address meniscus injuries each year.