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.
The James Hogg iCAPTURE Centre for Cardiovascular and Pulmonary Research, Department of Pathology and Laboratory Medicine, University of British Columbia, Vancouver, British Columbia, Canada.
Jonathan C
JC
Choy
Vivian H Y
VH
Hung
Arwen L
AL
Hunter
Paul K
PK
Cheung
Bruce
B
Motyka
Ing Swie
IS
Goping
Tracy
T
Sawchuk
R Chris
RC
Bleackley
Thomas J
TJ
Podor
Bruce M
BM
McManus
David J
DJ
Granville
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