|Gayle E. Nugent-Derfus, now an engineer at Genentech, and UCSD bioengineering professor Robert Sah showed that shear forces on cartilage prompt chondrocyte cells to produce proteoglycan 4, a molecule that lubricates and protects joint surfaces. High-res photo.|
The results reported in Osteoarthritis and Cartilage are important in the ongoing efforts of the group led by Robert Sah, a Howard Hughes Medical Institute (HHMI) professor at UCSD’s Jacobs School of Engineering, to grow cartilage in the laboratory that can be used to replace patients’ injured or diseased joint surfaces.
“We have shown that shear forces on cartilage prompt chondrocyte cells in it to produce proteoglycan 4,” said Sah. “This is an important step toward our goal of eventually growing joint tissue for transplantation.”
Proteoglycan, a name that reflects its protein and polysaccharide components, is a basic building block of connective tissue throughout the body. The chondrocyte cells of cartilage make several forms of proteoglycan, including several that build up in cartilage and contribute to its stiffness. However, proteoglycan-4 is primarily secreted into the joint fluid where it coats and lubricates cartilage surfaces.
In a series of experiments, Sah’s team attached bovine stifle joints, which are similar to human knee joints, to a bio-reactor that provided continuous irrigation with sterile nutritional fluids under normal physiological conditions. Immobile joints were compared to joints that were flexed 24 hours in a way that mimicked walking motions. The flexing was provided by a specially designed continuous passive motion device.
The team measured up to a three-fold increase in chondrocytes secreting proteoglycan 4 in continuously flexed joints compared to immobile controls. The flexing motion caused cartilage on the surfaces of opposing bones to slide against each other, creating so-called shear forces. In one large surface region of continuously sliding cartilage, 40 percent of the chondrocytes were secreting proteoglycan 4, whereas in the same areas of cartilage in immobilized joints only 13 percent of the chondrocytes were secreting proteoglycan 4. In areas of the joints exposed to only intermittent cartilage sliding, the effect on proteoglycan 4 production was intermediate between continuously sliding and immobilized regions of the joints.
“A challenge for us is to create large tissue grafts for transplantation,” said Sah. “We are systematically addressing the technical challenges to maintain and grow healthy fragments of bone and cartilage in the laboratory and now we can use nature’s self-regulating system, whereby application of shear forces to this tissue increases its synthesis of proteoglycan 4.”
Scientists have known for years that defects in a gene for proteoglycan 4 result in a type of childhood joint failure that resembles osteoarthritis in the elderly. Sah’s goal is to stimulate healthy chondrocytes in cartilage tissue grown in the laboratory to form robust tissue that makes proteoglycan 4 and has a smooth, well-lubricated surface.
|When the knee joint is not subjected to continuous passive motion (left), less proteoglycan 4 is produced by chondrocyte cells than when the knee joint is continuously moved (right). The dark purple stain is a specific visualization tool to identify chondrocyte cells in cartilage that are producing proteoglycan 4. Higher-resolution version.|
The paper in Osteoarthritis and Cartilage, which the journal will make available online the week of Dec. 4, was co-authored by Sah, Gayle E. Nugent-Derfus, now an engineer at Genentech, Inc., Dr. William D. Bugbee, associate adjunct professor of orthopaedics at UCSD and an orthopedic surgeon at Scripps Memorial Hospital in La Jolla, CA, and 13 researchers at UCSD, including six undergraduate students.
Sah’s research project and team were funded by the National Institutes of Health (NIH), the National Science Foundation (NSF), the Whitaker Foundation, Irwin and Joan Jacobs, and the UCSD Stein Institute for Research on Aging.
In addition, the project is benefiting from a $1million, four-year grant from HHMI to UCSD for Sah to develop innovative educational programs that "ignite the scientific spark in a new generation of students.” A central project accomplishment was made by undergraduates working under the supervision of Sah’s team members, especially engineers at Orthofix International’sVista, CA-based BREG, Inc. division. “Undergraduate seniors designed, built, and tested the prototype of the bioreactors in which the experiments were conducted,” said Sah. “Thanks to continuing collaboration with BREG and support from HHMI and NSF, additional undergraduates are now working on the project to improve the design of the bioreactor to enable the next round of research and development.”
G.E. Nugent-Derfus, T. Takara, J.K. O'Neill, S.B. Cahill, S. Görtz, T. Pong, H. Inoue, N.M. Aneloski, W.W. Wang, K.I. Vega, T.J. Klein, N.D. Hsieh-Bonassera, W.C. Bae, J.D. Burke, W.D. Bugbee, and R.L. Sah, "Continuous passive motion applied to whole joints stimulates chondrocyte biosynthesis of PRG4" (2006). Osteoarthritis and Cartilage.