Differentiation of chondrocytes, characterization of cell surface of chondrocytes and deleterious effects of MRI on cartilage repair
Date Issued
2007
Date
2007
Author(s)
Hsieh, Chang-Hsun
DOI
en-US
Abstract
Background: The proliferation and self-repair of articular cartilage are both poor. In fact, a number of age-related changes occur in the articular cartilage, including the free radical increase, a decrease in the number of chondrocytes and an increase in the degradation of matrix components. Such an age-related degeneration of articular cartilage could be a major risk factor for the development of osteoarthritis. Therefore, a lot of medical researches focused on the characters of chondrocytes in tissue engineering and clinic research. For instance, (1) As the MRI is a non-invasive physical assessment method, it is prevailing in clinic, including brain CT, tumor tracing, and assessment of surgery recovery…etc. In recent years, many papers discuss about the influence of high magnetic field or radiation on the cell, physiology and psychology, including expression of c-jun and c-fos genes, Ca2+ flux change, particularly Ca2+ entry from the extracellular environment through the plasma membrane, and induction of cloustrophobia syndrome…etc. Studies with cellular systems using different exposure setups, exposure durations, amplitudes, frequencies and wave forms indicate that biological effects of magnetic fields on cellular systems are at hand, but there is no definite conclusion in this field. In the first aim, we want to monitor the effect of the high magnetic field on human chondrocytes. (2) Atomic force microscope (AFM) permits the characterization of biologic samples ranging from a single molecules to whole cells (nanometers to micrometers), it is a powerful tool for exploring the shape of a single cell, the properties of a cellular membrane, or the interaction of intermolecular forces, such as adhesion force and stiffness. The second aim of this study is to use AFM technique to differentiate the mechanical behavior of the chondrocytes between the young modulus (normal) and the old modulus (OA). (3) Mesenchymal stem cells have multi-potential differentiation capacities, such as fat, muscle, bone and cartilage. In addition, primary culture of chondrocytes tend to lose the chondro-property and de-differentiation in monolayer culture condition during subculture. The third aim is to evaluate the characterization of MSCs differential capacities and establishing the 3D culture system for chondrocytes in vitro.
Method: (1) The 3T magnetic field was provided by the machine of Magnetic Resonance Imaging. The human chondrocytes were directly exposed to a 3-tesla (T) magnetic field (MF group) or a 3-T static magnetic field plus 125.3 MHz radio frequency (MF+RF group), and cell proliferation, apoptosis, cytosolic Ca2+ ([Ca2+]i) fluxes and expression of the apoptosis-related proteins of the treated cells were examined to assess the effects of the treatments. In the pig study, we examined the effects of the treatments on the recovery of surgically damaged pig knees. (2) We used AFM to observe a single chondrocyte cell directly and measured the dimensions of the cells. In addition, the receptors of cellular membrane were monitored by FACS. (3) The MSCs were isolated from human bone marrow. The MSCs were induced to adipogenesis, myogenesis, osteogenesis and chondrogenesis separately by respective inducers. The differentiated cell lineages were confirmed by specific markers expression by immunochemical staining. In addition, the RT-PCR system was applied for monitoring the property of chondrocytes in monolayer culture condition during subculture. Besides, chondrocytes which were seeded in the PGA/PLA scaffold was evaluated by using SEM observation.
Results and discussions: (1) A 3-T static magnetic field and radio frequency suppressed cell growth and induced apoptosis through p53, p21, p27 and Bax protein expression. In the pig model, we found that MRI surveillance had a deleterious effect on the recovery of the damaged knee cartilage. Magnetic strength, with or without concurrent radio frequency, suppressed chondrocyte growth in vitro and affected recovery of damaged knee cartilage in vivo in the pig model. The possible reason is chondrocytes exposing to the high magnetic field inducing cell apoptosis or cell transformation in the cartilage repair process. (2) The AFM revealed differences in the sizes and structures between the young modulus (normal) and the old modulus (OA). These findings suggested that the mechanical properties of normal chondrocytes substantially differed from those of OA chondrocytes. This new approach could be a useful technique for investigating age-related changes in the properties of human chondrocytes. (3) The MSCs from human bone marrow could differentiate into adipogenesis, myogenesis, osteogenesis and chondrogenesis processes. The morphology of human chondrocytes can be maintained after six passages both in petri dish and on scaffold but the cells were easily to be transformed to fibroblasts after eight passages. The scaffold laminated by PLA-coated PGA fiber was suitable for chondrocytes growth and showed better cell attachment under SEM investigation.
Subjects
chondrocytes, MRI
AFM
adhesion force
stiffness
MSCs and PLA/PGA
Type
other