TY - JOUR
T1 - Inhibition of cytotoxic T-lymphocyte-triggered apoptosis by target cell surface-coupled aprotinin
AU - Wagner, Ludwig
AU - Avery, Robin K.
AU - Bensinger, Lisa
AU - Kusinitz, Felicity
AU - Hibberd, Patricia L.
AU - Pasternack, Mark S.
N1 - Funding Information:
Acknowledgements-This work was supported by the American Cancer Society (IM 671) and by the National Institutes of Health (CA 45658).D r Ludwig Wagner was the recipient of a Max Kade Postdoctoral Research Exchange Grant and Dr Robin Avery was the recipient of a postdoctoral research fellowship from the American Cancer Society. We thank Dr Michael Bigby for his guidance regarding the use of the FAC-Scan flow cytometer.
PY - 1995/8
Y1 - 1995/8
N2 - A variety of recent investigations have implicated granzymes A and/or B in the target cell nuclear injury which accompanies cytotoxic T-lymphocyte-mediated cytolysis. Since soluble antiproteases have has limited efficacy in inhibiting CTL-mediated lysis, we developed a method to couple aprotinin, a peptide inhibitor of serine proteases, to the surface of target cells. Aprotinin modified by N-succinimidyl 3-(2-pyridyldithio)propionate retained trypsin-inhibitory activity, and target cells modified with aprotinin had demonstrable cell surface trypsin-inhibitory activity. Flow cytometry demonstrated that aprotinin was detectable on the target cell surface but underwent modulation at a rather rapid rate. When radiolabeled, aprotinin-coupled target cells were studied in 1-2 hr CTL assays, 51Cr release was little affected, but 125IUdR release was reduced up to 75% compared to controls. Corresponding apoptosis analysed by agarose gel electrophoresis and direct cytologic visualization was similarly reduced. Thus, aprotinin bound to the surface of target cells selectively protected target cells against CTL-mediated nuclear injury, and may serve as a model for the development of novel inhibitors of CTL-mediated lysis.
AB - A variety of recent investigations have implicated granzymes A and/or B in the target cell nuclear injury which accompanies cytotoxic T-lymphocyte-mediated cytolysis. Since soluble antiproteases have has limited efficacy in inhibiting CTL-mediated lysis, we developed a method to couple aprotinin, a peptide inhibitor of serine proteases, to the surface of target cells. Aprotinin modified by N-succinimidyl 3-(2-pyridyldithio)propionate retained trypsin-inhibitory activity, and target cells modified with aprotinin had demonstrable cell surface trypsin-inhibitory activity. Flow cytometry demonstrated that aprotinin was detectable on the target cell surface but underwent modulation at a rather rapid rate. When radiolabeled, aprotinin-coupled target cells were studied in 1-2 hr CTL assays, 51Cr release was little affected, but 125IUdR release was reduced up to 75% compared to controls. Corresponding apoptosis analysed by agarose gel electrophoresis and direct cytologic visualization was similarly reduced. Thus, aprotinin bound to the surface of target cells selectively protected target cells against CTL-mediated nuclear injury, and may serve as a model for the development of novel inhibitors of CTL-mediated lysis.
KW - apoptosis
KW - aprotinin
KW - cell-mediated cytotoxicity
KW - cytotoxic T-lymphocyte
KW - granzyme A
KW - protease inhibition
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U2 - 10.1016/0161-5890(95)00054-I
DO - 10.1016/0161-5890(95)00054-I
M3 - Article
C2 - 7565812
AN - SCOPUS:0029080301
SN - 0161-5890
VL - 32
SP - 853
EP - 864
JO - Molecular Immunology
JF - Molecular Immunology
IS - 12
ER -