The effects of serine proteinase inhibitors on bone resorption in vitro

• Published in: Journal of endocrinology Vol. 178; no. 3; pp. 437 - 447
• Main Authors: Tumber, A, Papaioannou, S, Breckon, J, Meikle, MC, Reynolds, JJ, Hill, PA
• Format: Journal Article
• Language: English
• Published: Colchester BioScientifica 01-09-2003; Portland Press
• Subjects: alpha-2-Antiplasmin - pharmacology; Amyloid beta-Protein Precursor; Animals; Animals, Newborn; Aprotinin - pharmacology; Biological and medical sciences; Bone and Bones - drug effects; Bone and Bones - physiology; Bone Marrow Cells; Bone Resorption - metabolism; Carrier Proteins - genetics; Cells, Cultured; Culture Techniques; Dose-Response Relationship, Drug; Fundamental and applied biological sciences. Psychology; Gene Expression; Mice; Osteoblasts - drug effects; Osteoblasts - metabolism; Osteoclasts - drug effects; Osteoclasts - metabolism; Plasminogen Activator Inhibitor 1 - genetics; Plasminogen Activator Inhibitor 2 - genetics; Protease Nexins; Receptors, Cell Surface - genetics; Receptors, Urokinase Plasminogen Activator; Reverse Transcriptase Polymerase Chain Reaction; Serine Proteinase Inhibitors - pharmacology; Skeleton and joints; Tissue Plasminogen Activator - genetics; Urokinase-Type Plasminogen Activator - genetics; Vertebrates: osteoarticular system, musculoskeletal system; Serine; Rodentia; Resorption; Osteoclast; Gene expression; In vitro; Plasminogen plasmin system; Osteoarticular system; Vertebrata; Mammalia; Cell line; Mouse; Osteoblast; Bone; Protease inhibitor
• ISSN: 0022-0795; 1479-6805
• DOI: 10.1677/joe.0.1780437

The aims of this study were to identify the role and sites of action of serine proteinases (SPs) in bone resorption, a process which involves a cascade of events, the central step of which is the removal of bone matrix by osteoclasts (OCs). This resorbing activity, however, is also determined by recruitment of new OCs to future resorption sites and removal of the osteoid layer by osteoblasts (OBs), which enables OCs to gain access to the underlying mineralized bone. The resorption systems we have studied consisted of (i) neonatal calvarial explants, (ii) isolated OCs cultured on ivory slices, (iii) mouse OBs cultured on either radiolabelled type I collagen films or bone-like matrix, (iv) bone marrow cultures to assess OC formation and (v) 17-day-old fetal mouse metatarsal bone rudiments to assess OC migration and fusion. Two separate SP inhibitors, aprotinin and alpha(2)-antiplasmin dose-dependently inhibited (45)Ca release from neonatal calvarial explants: aprotinin (10(-6) M) was the most effective SP inhibitor, producing a maximum inhibitory effect of 55.9%.Neither of the SP inhibitors influenced either OC formation or OC resorptive activity. In contrast, each SP inhibitor dose-dependently inhibited OB-mediated degradation of both type I collagen fibrils and non-mineralized bone matrix. In 17-day-old metatarsal explants aprotinin produced a 55% reduction in the migration of OCs from the periosteum to the mineralized matrix after 3 days in culture but after 6 days in culture aprotinin was without effect on OC migration. Primary mouse osteoblasts expressed mRNA for urokinase type plasminogen activator (uPA), tIssue type plasminogen activator (tPA), the type I receptor for uPA, plasminogen activator inhibitor types I and II and the broad spectrum serine proteinase inhibitor, protease nexin I. In situ hybridization demonstrated expression of tPA and uPA in osteoclasts disaggregated from 6-day-old mouse long bones. We propose that the regulation of these various enzyme systems within bone tIssue determines the sites where bone resorption will be initiated.