Proteolytic activity of Triatoma infestans saliva associated with PAR-2 activation and vasodilation

Author(s):Show less -

Karla A. Oliveira ^[1] ; Ricardo J. S. Torquato ^[2] ; Daniela C. G. Garcia Lustosa ^[3] ; Tales Ribeiro ^[4] ; Bruno W. L. Nascimento ^[5] ; Lilian C. G. de Oliveira ^[6] ; Maria A. Juliano ^[7] ; Thaysa Paschoalin ^[8] ; Virginia S. Lemos ^[9] ; Ricardo N. Araujo ; Marcos H. Pereira ; Aparecida S. Tanaka

Affiliation:Show less -

^[1] Federal University of Piauí. Department of Biochemistry and Pharmacology - Brasil

^[2] Federal University of São Paulo. Department of Biochemistry - Brasil

^[3] Federal University of Minas Gerais. Institute of Biomedical Sciences. Department of Pharmacology - Brasil

^[4] Federal University of Minas Gerais. Institute of Biomedical Sciences. Department of Parasitology - Brasil

^[5] Federal University of Minas Gerais. Institute of Biomedical Sciences. Department of Parasitology - Brasil

^[6] Federal University of São Paulo. National Institute of Pharmacology and Molecular Biology. Department of Biophysics - Brasil

^[7] Federal University of São Paulo. National Institute of Pharmacology and Molecular Biology. Department of Biophysics - Brasil

^[8] Federal University of São Paulo. National Institute of Pharmacology and Molecular Biology. Department of Biophysics - Brasil

^[9] Federal University of Minas Gerais. Institute of Biomedical Sciences. Department of Physiology and Biophysics - Brasil

• Document type: Journal article
• Source: Journal of Venomous Animals and Toxins including Tropical Diseases; v. 27, 2021-03-08.

Abstract

ABSTRACT Background Triatoma infestans (Hemiptera: Reduviidae) is a hematophagous insect and the main vector of Trypanosoma cruzi (Kinetoplastida: Trypanosomatidae). In the present study, the authors investigated whether a serine protease activity from the saliva of T. infestans has a role in vasomotor modulation, and in the insect-blood feeding by cleaving and activating protease-activated receptors (PARs). Methods T. infestans saliva was chromatographed as previously reported for purification of triapsin, a serine protease. The cleavage activity of triapsin on PAR peptides was investigated based on FRET technology. Mass spectrometry was used to analyze the sites of PAR-2 peptide cleaved by triapsin. NO measurements were performed using the DAN assay (2,3-diaminonapthalene). The vasorelaxant activity of triapsin was measured in vessels with or without functional endothelium pre-contracted with phenylephrine (3 µM). Intravital microscopy was used to assess the effect of triapsin on mouse skin microcirculation. Results Triapsin was able to induce hydrolysis of PAR peptides and showed a higher preference for cleavage of the PAR-2 peptide. Analysis by mass spectrometry confirmed a single cleavage site, which corresponds to the activation site of the PAR-2 receptor. Triapsin induced dose-dependent NO release in cultured human umbilical vein endothelial cells (HUVECs), reaching a maximum effect at 17.58 nM. Triapsin purified by gel-filtration chromatography (10-16 to 10-9 M) was applied cumulatively to mouse mesenteric artery rings and showed a potent endothelium-dependent vasodilator effect (EC30 = 10-12 M). Nitric oxide seems to be partially responsible for this vasodilator effect because L-NAME (L-NG-nitroarginine methyl ester 300 µM), a nitric oxide synthetase inhibitor, did not abrogate the vasodilation activated by triapsin. Anti-PAR-2 antibody completely inhibited vasodilation observed in the presence of triapsin activity. Triapsin activity also induced an increase in the mouse ear venular diameter. Conclusion Data from this study suggest a plausible association between triapsin activity mediated PAR-2 activation and vasodilation caused by T. infestans saliva. (AU)

• FAPESP's process: 12/03657-8 - Inhibitor and proteases of ectoparasites: relationship of structure-function and identification of the role of these molecules in the interaction of diseases vector e their etiological agents
• Grantee: Aparecida Sadae Tanaka
• Support Opportunities: Research Projects - Thematic Grants