student research day - Case Western Reserve University School of ...

Rebekah C. Brown
Proteomic Analysis of Human Synovial Fluid, Synovium and Cartilage in Healthy and Osteoarthritic Subjects: An
Investigation of the Knee Joint
Rebekah C. Brown, Reuben Gobezie MD., James Crish Ph.D., Eldra Daniels, Eric Rodriguez, Mark Chance Ph.D.,
Gurkan Bebek Ph.D., Tim Henderson, Serguei Ilchenko Ph.D., Giri Gokulrandan Ph.D., Patrick Leahy Ph.D. and
Chunbiao Li
University Hospital Department of Orthopedics, Case Center for Proteomics and Bioinformatics, The Gene
Expression and Genotyping Core Facility, Case Comprehensive Cancer Center, Case Western Reserve University
Osteoarthritis is a multifactorial degenerative joint disease characterized by pathophysiologic changes to
synovial joints. (e-medicine) Osteoarthritis (OA) of the knee joint is a huge problem affecting,
approximately, 27% of people over the age of 45. [MD Consult Epidemiology of Osteoarthritis
Rheumatic Diseases Clinics of North America - Volume 34, Issue 3 (August 2008)] The question to
ask is how do we mitigate OA and its effects? Through proteomics, “[t]he study of the structure and
function of proteins, including the way they work and interact with each other inside cells.”
[www.cancer.gov], we can answer key questions; namely, what is the proteome of an arthritic
individual’s synovium, synovial fluid, and cartilage; what are the biological pathway alterations between
these tissues and disease states; and what is the molecular level of alteration(s) defining osteoarthritis?
These questions were explored using several proteomic techniques and microarray analysis. The
synovial fluid, synovium and cartilage samples were prepared from five individuals for 1-D PAGE
electrophoresis with subsequent in-gel digestion and Western Blot analysis. The in-gel digestion
products were analyzed by FT/LTQ and OrbiTRAP mass spectral analysis, MASCOT and MASRAN
(MASCOT Results Analyzer) within the Case Center for Proteomics and Bioinformatics. Western Blot
analysis was performed targeting the proteins Gelsolin and Afamin. RNA isolation procedures were
performed in the Gobezie lab for microarray analysis at The Gene Expression and Genotyping Core
Facility, Case Comprehensive Cancer Center. Analysis showed that there are proteomic differences
between disease states. In synovial fluid, the proteome of the early osteoarthritis (EOA) group is greater
than the healthy group, which was greater than the late osteoarthritis (LOA) group. The LOA proteome is
greater than the EOA proteome in synovial tissue and cartilage. Additionally, within the same individual,
the proteome differs between tissue types. The proteome is larger in synovium versus that of synovial
fluid and cartilage. Preliminary microarray analysis results substantiate these proteomic results. Results
are still pending regarding the biological pathway analysis. So far, it is evident that there are discrete
protein alterations between healthy and osteoarthritic individuals. This knowledge of proteomic changes
associated with OA can be translated into obtaining definitive diagnoses and earlier detection of the
disease. Focusing on a proteomic approach to study osteoarthritis, such as biological pathway
alteration, can potentially yield very insightful information about the arthritic process.
Supported by NIH Heart, Lung and Blood Institute Grant: Ruth L. Kirschstein National Research Service Award
Short-Term Institutional Research Training Grants (T35)
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