Immunolocalisation of BPTI-like serine proteinase inhibitory proteins in mast cells, chondrocytes and intervertebral disc fibrochondrocytes of ovine and bovine connective tissues. An immunohistochemical and biochemical study.
A polyclonal anti-bovine pancreatic trypsin inhibitor (BPTI) IgY was raised in chickens immunised with aprotinin. The anti-BPTI IgY was subsequently isolated from egg yolks and purified to homogeneity by affinity chromatography on immobilised aprotinin and by Superose 6 size exclusion fast protein liquid chromatography (FPLC). Immunoblotting with the chicken IgY demonstrated its specificity for BPTI; 3.9 ng BPTI could be detected by this technique. There was no crossreactivity against alpha1-proteinase inhibitor (human and sheep), inter-alpha-trypsin inhibitor (human and sheep), secretory leucocyte proteinase inhibitor or a range of serine proteinase inhibitory proteins (SPIs) isolated from plant sources (soybean and lima bean trypsin inhibitor, potato trypsin and chymotrypsin inhibitors) or serum SPIs (antithrombin-III, alpha2-macroglobulin). Immunoblotting using the anti-BPTI IgY identified the 6- to 12- and 58-kDa forms of endogenous ovine cartilage SPIs in cartilage extracts, confirming the interrelationship of the ovine cartilage SPIs with BPTI. BPTI-domain SPIs were immunolocalised within mast cells of ovine and bovine duodenum, lung and pancreas, and in ovine and bovine bronchial cartilage chondrocytes, chondrocytes of the superficial and intermediate zones of articular cartilage and in the fibrochondrocytes/chondrocytes of the nucleus
Melrose
J
J
The Institute of Bone and Joint Research, The Royal North Shore Hospital of Sydney, and Department of Surgery of The University of Sydney, St. Leonards, NSW, Australia. jmelrose@mail.usyd.edu.au
Smith
S
S
Rodgers
K
K
Little
C
C
Burkhardt
D
D
Ghosh
P
P
D000818:Animals; D007611:Aprotinin / Q000032:analysis* / Q000276:immunology; D002417:Cattle; D019902:Chondrocytes / Q000737:chemistry*; D003238:Connective Tissue / Q000737:chemistry; D015151:Immunoblotting / Q000379:methods; D007403:Intervertebral Disc / Q000737:chemistry*; D008407:Mast Cells / Q000737:chemistry*; D015842:Serine Proteinase Inhibitors / Q000032:analysis* / Q000276:immunology; D012756:Sheep; D014361:Trypsin Inhibitors / Q000032:analysis* / Q000276:immunology
D015842:Serine Proteinase Inhibitors; D014361:Trypsin Inhibitors; D007611:Aprotinin