Granzyme B induces smooth muscle cell apoptosis in the absence of perforin: involvement of extracellular matrix degradation.
OBJECTIVE
T cell-induced cytotoxicity, of which granzyme B is a key mediator, is believed to contribute to the pathogenesis of inflammatory vascular diseases. In this report, we investigate the mechanism of granzyme B-induced smooth muscle cell (SMC) death.


RESULTS
The addition of purified granzyme B alone to cultured SMCs caused a significant reduction in cell viability. Chromatin condensation, phosphatidylserine externalization, and membrane blebbing were observed, indicating that the mechanism of granzyme B-induced SMC death was through apoptosis. Activated splenocytes from perforin-knockout mice induced SMC death through a granzyme B-mediated pathway. Inhibition of the proteolytic activities of caspases and granzyme B prevented granzyme B-induced SMC death, whereas attenuation of granzyme B internalization with mannose-6-phosphate (M6P) did not. Further, granzyme B induced the cleavage of several SMC extracellular proteins, including fibronectin, and reduced focal adhesion kinase phosphorylation.


CONCLUSIONS
These results indicate that granzyme B can induce apoptosis of SMCs in the absence of perforin by cleaving extracellular proteins, such as fibronectin.
Choy
Jonathan C
JC
The James Hogg iCAPTURE Centre for Cardiovascular and Pulmonary Research, Department of Pathology and Laboratory Medicine, University of British Columbia, Vancouver, British Columbia, Canada.
Hung
Vivian H Y
VH
Hunter
Arwen L
AL
Cheung
Paul K
PK
Motyka
Bruce
B
Goping
Ing Swie
IS
Sawchuk
Tracy
T
Bleackley
R Chris
RC
Podor
Thomas J
TJ
McManus
Bruce M
BM
Granville
David J
DJ
D000818:Animals; D017209:Apoptosis / Q000502:physiology*; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000502:physiology; D003331:Coronary Vessels / Q000201:enzymology / Q000378:metabolism; D005109:Extracellular Matrix / Q000201:enzymology / Q000378:metabolism*; D016326:Extracellular Matrix Proteins / Q000378:metabolism; D053804:Granzymes; D006367:HeLa Cells / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D008562:Membrane Glycoproteins / Q000172:deficiency* / Q000502:physiology; D051379:Mice; D008810:Mice, Inbred C57BL; D009131:Muscle, Smooth, Vascular / Q000201:enzymology / Q000502:physiology; D032389:Myocytes, Smooth Muscle / Q000201:enzymology / Q000502:physiology*; D054353:Perforin; D052899:Pore Forming Cytotoxic Proteins; D051381:Rats; D012697:Serine Endopeptidases / Q000378:metabolism / Q000502:physiology*; D013154:Spleen / Q000166:cytology / Q000378:metabolism
D016326:Extracellular Matrix Proteins; D008562:Membrane Glycoproteins; D052899:Pore Forming Cytotoxic Proteins; D054353:Perforin; C509213:GZMB protein, human; D053804:Granzymes; C509214:Gzmb protein, mouse; C509215:Gzmb protein, rat; D012697:Serine Endopeptidases; C505407:CASP3 protein, human; C505409:Casp3 protein, mouse; C505410:Casp3 protein, rat; D053148:Caspase 3; D020169:Caspases