The protein disulphide isomerase inhibitor CxxCpep modulates oxidative burst and mitochondrial function in platelets

Author(s)

Gaspar, Renato S. ^{[1, 2, 3]} ; Mansilla, Santiago ^{[4, 5]} ; Vieira, Victor A. ^{[2, 6]} ; Silva, Ludmila B. da ^{[2, 7]} ; Gibbins, Jonathan M. ^[1] ; Castro, Laura ^{[4, 6]} ; Trostchansky, Andres ^{[4, 6]} ; Paes, Antonio Marcus de A. ^{[2, 7]}

Affiliation

^[1] Univ Reading, Sch Biol Sci, Inst Cardiovasc & Metab Res, Reading, Berks - England

^[2] Univ Fed Maranhao, Biol & Hlth Sci Ctr, Dept Physiol, Lab Expt Physiol, Sao Luis, Maranhao - Brazil

^[3] Univ Sao Paulo, Sch Med, Hlth Inst InCor, Lab Vasc Biol, Sao Paulo - Brazil

^[4] Univ Republica, Fac Med, Dept Bioquim, Montevideo - Uruguay

^[5] Univ Republica, Fac Med, Dept Metodos Cuantitat, Montevideo - Uruguay

^[6] Univ Republica, Fac Med, Ctr Invest Biomed CEINBIO, Montevideo - Uruguay

^[7] Univ Fed Maranhao, Hlth Sci Grad Program, Biol & Hlth Sci Ctr, Sao Luis, Maranhao - Brazil

• Document type: Journal article
• Source: Free Radical Biology and Medicine; v. 172, p. 668-674, AUG 20 2021.
• Web of Science Citations: 0

Abstract

Background: We have previously described CxxCpep, a peptide with anti-platelet properties that inhibits peri/ epicellular protein disulphide isomerase (pecPDI) by forming a mixed disulfide bond with Cys400 within the pecPDI active site. Objectives: Here we sought to determine if pecPDI targeted by CxxCpep is relevant to redox mechanisms downstream of the collagen receptor GPVI in platelets. Methods and results: Restriction of effects of CxxCpep to the platelet surface was confirmed by LC-MS/MS following cell fractionation. Platelet aggregation was measured in platelet-rich plasma (PRP) incubated with 30 mu M CxxCpep or vehicle. CxxCpep inhibited collagen-induced platelet aggregation but exerted no effect in TRAP-6-stimulated platelets. PRP was incubated with DCFDA to measure oxidative burst upon platelet adhesion to collagen. Results showed that CxxCpep decreased oxidative burst in platelets adhered to immobilized collagen while the number of adherent cells was unaffected. Furthermore, flow cytometry studies using a FITC-maleimide showed that the GPVI agonist CRP stimulated an increase in free thiols on the platelet outer membrane, which was inhibited by CxxCpep. Finally, CxxCpep inhibited platelet mitochondrial respiration upon activation with collagen, but not with thrombin. Conclusions: Our data suggest that pecPDI is a potential modulator of GPVI-mediated redox regulation mechanisms and that CxxCpep can be further exploited as a template for new antiplatelet compounds. (AU)

• FAPESP's process: 20/15944-8 - Investigation of peri/epicellular protein disulphide isomerase as a novel regulator of the platelet-endothelium interaction in normoglycaemia and Diabetes
• Grantee: Renato Simões Gaspar
• Support Opportunities: Scholarships in Brazil - Post-Doctoral