ADAM15 exerts an antiapoptotic effect on osteoarthritic chondrocytes via up-regulation of the X-linked inhibitor of apoptosis.
OBJECTIVE
To investigate the capacity of ADAM15, a disintegrin metalloproteinase that is up-regulated in osteoarthritic (OA) cartilage, to protect chondrocytes against apoptosis induced by growth factor deprivation and genotoxic stress.


METHODS
Caspase 3/7 activity was determined in primary OA and ADAM15-transfected T/C28a4 chondrocytes upon exposure to the DNA-damaging agent camptothecin or serum withdrawal. Camptothecin-induced cytotoxicity was determined by measuring cellular ATP content. (Anti-)apoptotic proteins were analyzed by immunoblotting, and levels of messenger RNA (mRNA) for X-linked inhibitor of apoptosis (XIAP) were determined using real-time polymerase chain reaction. RNA interference was applied for down-regulation of ADAM15 and XIAP expression. Immunohistochemistry analysis of normal and OA cartilage samples was performed using XIAP- and ADAM15-specific antibodies.


RESULTS
ADAM15-transfected chondrocytes cultured on a collagen matrix displayed significantly reduced caspase 3/7 activity upon serum or intermittent matrix withdrawal, compared with vector-transfected control cells. Apoptosis induction by camptothecin exposure also led to significantly elevated caspase 3/7 activity and reduced cell viability of the vector-transfected compared with ADAM15-transfected chondrocytes. Increased levels of activated caspase 3 and cleaved poly(ADP-ribose) polymerase were detected in the vector controls. XIAP, an inhibitor of activated caspase 3, was significantly up-regulated ( approximately 3-fold) at the protein and mRNA levels in ADAM15-transfected chondrocytes upon camptothecin treatment. Specific down-regulation of either ADAM15 or XIAP in OA chondrocytes led to significant sensitization to camptothecin-induced caspase 3/7 activity. Immunohistochemical analysis revealed low to moderate XIAP expression in normal specimens and markedly increased XIAP staining, colocalizing with ADAM15, in OA cartilage.


CONCLUSIONS
ADAM15 conveys antiapoptotic properties to OA chondrocytes that might sustain their potential to better resist the influence of death-inducing stimuli under pathophysiologic conditions.
Böhm
Beate
B
Johann Wolfgang Goethe University, Frankfurt am Main, Germany.
Hess
Sandra
S
Krause
Kristin
K
Schirner
Andrea
A
Ewald
Werner
W
Aigner
Thomas
T
Burkhardt
Harald
H
D051722:ADAM Proteins / Q000235:genetics / Q000378:metabolism*; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D001798:Blood Proteins / Q000494:pharmacology; D002166:Camptothecin / Q000494:pharmacology; D002352:Carrier Proteins / Q000494:pharmacology; D053148:Caspase 3 / Q000235:genetics / Q000378:metabolism; D053179:Caspase 7 / Q000235:genetics / Q000378:metabolism; D002478:Cells, Cultured; D019902:Chondrocytes / Q000166:cytology* / Q000187:drug effects / Q000502:physiology; D015536:Down-Regulation / Q000502:physiology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D010003:Osteoarthritis / Q000378:metabolism* / Q000473:pathology*; D012333:RNA, Messenger / Q000378:metabolism; D034741:RNA, Small Interfering; D013312:Stress, Physiological / Q000502:physiology; D014162:Transfection; D015854:Up-Regulation / Q000187:drug effects / Q000502:physiology; D051636:X-Linked Inhibitor of Apoptosis Protein / Q000235:genetics / Q000378:metabolism*
D001798:Blood Proteins; D002352:Carrier Proteins; D004791:Enzyme Inhibitors; C489454:Fgfbp1 protein, rat; D036341:Intercellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D051636:X-Linked Inhibitor of Apoptosis Protein; C496866:XIAP protein, human; C505407:CASP3 protein, human; C505520:CASP7 protein, human; D053148:Caspase 3; D053179:Caspase 7; D051722:ADAM Proteins; C098660:ADAM15 protein, human; D002166:Camptothecin