Superficial zone chondrocytes in normal and osteoarthritic human articular cartilages synthesize novel truncated forms of inter-alpha-trypsin inhibitor heavy chains which are attached to a chondroitin sulfate proteoglycan other than bikunin.
OBJECTIVE
We have examined the occurrence of the inflammation-associated inter-alpha-trypsin inhibitor (IalphaI) components, bikunin, heavy chain (HC)1 and HC2 in normal cartilage and osteoarthritis (OA) cartilage and synovial fluids.


METHODS
Cartilage extracts from normal donors and late-stage OA patients, and synovial fluids from OA patients were studied by Western blot with multiple antibodies to bikunin, HC1 and HC2. Cell and matrix localization was determined by immunohistochemistry and mRNA by RT-PCR.


RESULTS
Bikunin.chondroitin sulfate (CS) and IalphaI were abundant in OA cartilages, but virtually undetectable in normal. In both OA and normal cartilages, HCs were largely present in a novel C-terminally truncated 50-kDa form, with most, if not all of these being attached to CS on a proteoglycan other than bikunin. Synovial fluids from OA patients contained bikunin.CS and full-length (approximately 90 kDa) HCs linked to hyaluronan (HA) as HC.HA (SHAP.HA). Immunohistochemistry showed intracellular and cell-associated staining for bikunin and HCs, consistent with their synthesis by superficial zone chondrocytes. PCR on multiple human normal and OA cartilage samples detected transcripts for HC1 and HC2 but not for bikunin. In OA cartilages, immunostaining was predominantly matrix-associated, being most intense in regions with a pannus-like fibrotic overgrowth.


CONCLUSIONS
The truncated structure of HCs, their attachment to a proteoglycan other than bikunin, PCR data and intracellular staining are all consistent with synthesis of HC1 and HC2 by human articular chondrocytes. The presence of bikunin.CS and IalphaI in OA cartilage, but not in normal, appears to be due to diffusional uptake and retention through fibrillated (but not deeply fissured) cartilage surfaces.
Yoshihara
Y
Y
Department of Biochemistry, Rush University Medical Center, Chicago, IL, USA.
Plaas
A
A
Osborn
B
B
Margulis
A
A
Nelson
F
F
Stewart
M
M
Rugg
M S
MS
Milner
C M
CM
Day
A J
AJ
Nemoto
K
K
Sandy
J D
JD
D000510:Alpha-Globulins / Q000096:biosynthesis* / Q000737:chemistry; D015153:Blotting, Western; D002358:Cartilage, Articular / Q000378:metabolism*; D019902:Chondrocytes / Q000378:metabolism*; D011508:Chondroitin Sulfate Proteoglycans / Q000378:metabolism; D002809:Chondroitin Sulfates / Q000737:chemistry; D006801:Humans; D006820:Hyaluronic Acid / Q000378:metabolism; D010003:Osteoarthritis / Q000378:metabolism*; D011509:Proteoglycans / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013582:Synovial Fluid
D000510:Alpha-Globulins; D011508:Chondroitin Sulfate Proteoglycans; D011509:Proteoglycans; C016001:alpha-1-microglobulin; C011877:inter-alpha-inhibitor; D006820:Hyaluronic Acid; D002809:Chondroitin Sulfates