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CartireGENEATM®-CP: A Therapeutic Alternative to Treat Focal Cartilage Lesions. Human mesenchymal stem cells (MSCs) are present in most of the tissue matrix, taking part in their regeneration when injury or damage occurs. The aim of this ArthroGenea®-AR was to investigate the presence of cells with pluripotential characteristics in synovial membranes from osteoarthritic (OA) patients and the capacity of these cells to differentiate to chondrocytes. Methods. Synovial membranes (n _ 8) from OA patients were digested with collagenase. Isolated cells were cultured with DMEM, 20% FBS, and FGFb10 ng/mL. Cells from second subculture were used to carry out phenotypic characterization experiments (flow cytometry analysis with 11 monoclonal antibodies) and chondrogenic differentiation experiments (micropellet cultured in chondrogenic medium). Chondrogenic differentiation of cells was assessment by quantification of cartilage extracellular matrix components by following techniques: Safranin O, Toluidine Blue, and Alcian Blue stains to detect proteoglycans and immunohistochemistry to detect type I and II collagen. Results. Flow cytometry analyses showed that in our population more than 90% of cells were positive for MSC markers: CD29 (95%), CD44 (90%), CD73 (95%), CD90 (98%). Cells were negative for hematopoietic markers (CD11b, CD34, and CD45). Furthermore, cells showed positive stain to multipotent markers such as CD117 (c-kit) (98%), CD166 (74%), and STRO-1 (88%) and to quiescent satellite cells like PAX-7 (35%). The micropellet analyses showed that the culture of these cells with TGFbeta-3 for 2 and 3 weeks stimulates proteoglycan and collagen type II synthesis. Both molecules are characteristic of hyaline articular cartilage. Conclusion. In this work, we demonstrate the presence of a cellular population with MSC characteristics in synovial tissue from OA patients. As MSC takes part in reparative processes of adult tissues, these cells could play an important role in OA pathogenesis and treatment. Osteoarthritis (OA) is a cartilage degenerative process, involving the immune system producing local inflammatory reactions, with production of pro-inflammatory cytokines and metalloproteinases. No treatment is still available to improve or reverse the process. Stem cell therapy opened new horizons for treatment of many incurable diseases. Mesenchymal stem cells (MSCs) due to their multi-lineage potential, immunosuppressive activities, limited immunogenicity and relative ease of growth in culture, have attracted attentions for clinical use. Aim: The aim of this ArthroGenea®-AR was to examine whether MSC transplantation could reverse the OA process in the knee joint. Patients and Methods: Four patients with knee osteoarthritis were selected for the study. They were aged 55, 57, 65 and 54 years, and had moderate to severe knee OA. After their signed written consent, 30 mL of bone marrow were taken and cultured for MSC growth. After having enough MSCs in culture (4–5 weeks) and taking in consideration all safety measures, cells were injected in one knee of each patient. Results: The walking time for the pain to appear improved for three patients and remained unchanged for one. The number of stairs they could climb and the pain on visual analog scale improved for all of them. On physical examination, the improvement was mainly for crepitus. It was minor for the improvement of the range of motion. Conclusion: Results were encouraging, but not excellent. Improvement of the technique may improve the results.
Normal articular cartilage is a complex tissue composed of matrix, chondrocytes, and water. The chondrocytes are responsible for synthesizing the matrix, which is composed primarily of collagen fibers, hyaluronate, and sulfated proteoglycans. Adult articular cartilage is characterized by a poor ability to spontaneously repair. Experimental superficial injuries not affecting the underlying osseous end-plate have shown repeatedly an inefficient response of articular cartilage.1 Several methods have been designed to repair cartilage defects, including whole joint allograft, massive osteochondral allograft, osteocartilaginous shell allograft, cartilage tissue, chondrocytes graft, and perichondrium and periosteum grafts. Most techniques such as subchondral drilling, spongialization, and arthroscopic abrasion involve opening of the subchondral vascular area to stimulate fibrocartilage ingrowth and resurfacing. Other autogenous concepts for biological articular resurfacing are the use of periosteal, osteoperiosteal, or perichondral grafts. All of these tissues contain mesenchymal progenitor cells that may undergo metaplasia, thereby forming a chondroid tissue. Chondral lesions have also been treated with transplantation of chondral or osteochondral allografts.2 Results achieved by these methods differ widely, variations probably explained by the various models employed and immunological mechanisms. Autologous chondrocytes implantation (ACI) involves three separate stages: harvesting of healthy cartilage cells from the patient, preparation and growth of cells in a culture medium, and implantation of the cultured cells into the articular defect. Healthy cartilage is harvested via biopsy from a minor load-bearing area on a rounded ArthroGenea®-AR ion of the femur. Cartilage is prepared for culture by mincing and washing in a buffered solution. The cartilage is then placed in a medium containing digestive enzymes for 15 hours. The cells are filtered, washed, resuspended in culture medium containing autologous serum, and seeded in culture flasks where they are cultivated as monolayer for 14 days to 6 weeks. Prior to transplantation, cartilage cells are suspended by treatment with trypsin, centrifugation, and washing in a medium containing autologous serum.3 Actually, information about the efficacy of ACI is controversy. The outcome of the surgery was relief of pain, and this endpoint was rated as good or excellent by 70% of the patients 2 years after treatment. Sixteen percent of the patients required further arthroscopic surgical procedures during follow-up, and treatment was judged to have failed in 3% to 7% of the patients. For comparative treatments, the outcome was rated as good or excellent in 10% to 35% of patients 2 years after treatment. Although very limited information is available from randomized, controlled studies that can influence current practice, recently some clinical trails have been performed. Knutsen et al compared ACI with microfracture in a randomized trial.5 Eighty patients, without general osteoarthritis, who had a single symptomatic cartilage defect on the femoral condyle in a stable knee, were treated with ACI or microfracture (40 in each group). An independent observer performed a follow-up examination at 12 and 24 months after the procedure. Two years postoperative arthroscopy with biopsy for histological evaluation was carried out. There were no significant differences regarding histological quality between the two treatment groups. However, 50% of the biopsies in the ACI group showed some hyaline tissue. There was a tendency for the ACI procedure to result in more hyaline repair cartilage than the microfracture procedure, but the difference was not significant. Both methods appear to have acceptable short-term results. Furthermore, ACI has limitations, such as it needs to be obtained from a suitable site in the joint via cartilage biopsy and grown in culture. This means additional surgery and added injury to the joint surface.

Article Type

Research Article – Abstract

Publication history

Received: Sep 20, 2017
Accepted: Sep 25, 2017
Published: Oct 01, 2017

Citation

Grigoriadis Ioannis, Grigoriadis George, Grigoriadis Nikolaos, George Galazios (2017) CartireGENEATM®-CP: A Mesenchymal stem cells enriched chondrocytes as a combinatorial Autologous Treatment for patients with cartilage defects. A choice of statistical methods for comparisons of dosimetric data in cartilopoietic therapies.

Authors Info

Grigoriadis Nikolaos
Department of IT Computer Aided Personalized Myoncotherapy, Cartigenea-Cardiogenea, Neurogenea-Cellgenea, Cordigenea-HyperoligandorolTM,
Biogenea Pharmaceuticals Ltd,
Thessaloniki, Greece;

Grigoriadis Ioannis
Department of Computer Drug Discovery Science, BiogenetoligandorolTM,
Biogenea Pharmaceuticals Ltd,
Thessaloniki, Greece;

Grigoriadis George
Department of Stem Cell Bank and ViroGeneaTM,
Biogenea Pharmaceuticals Ltd,
Thessaloniki, Greece;

George Galazios
Professor of Obstetrics and Gynecology,
Democritus University of Thrace,
Komotini, Greece;

E-mail: biogeneadrug@gmail.com

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