Insights into cathepsin-B activity in mature dentin matrix

Autor(es)

Carrilho, Marcela R. ^[1] ; Scaffa, Polliana ^[2] ; Oliveira, Vitor ^[3] ; Tjaderhane, Leo ^{[4, 5, 6]} ; Tersariol, Ivarne L. ^[7] ; Pashley, David H. ^[8] ; Tay, Franklin ^[8] ; Nascimento, Fabio D. ^[9]

Afiliação do(s) autor(es)

^[1] Midwestern Univ, Coll Dent Med Illinois, 555 31st St, SH 211-X, Downers Grove, IL 60515 - USA

^[2] Oregon Hlth & Sci Univ, Biomat & Biomech, Portland, OR 97201 - USA

^[3] Univ Fed Sao Paulo, Dept Biophys, Sao Paulo - Brazil

^[4] Univ Oulu, Oulu - Finland

^[5] Univ Helsinki, Helsinki Univ Hosp, Dept Oral & Maxillofacial Dis, Helsinki - Finland

^[6] Oulu Univ Hosp, Med Res Ctr Oulu MRC Oulu, Res Unit Oral Hlth Sci, Oulu - Finland

^[7] Univ Fed Sao Paulo, Dept Biochem, Sao Paulo - Brazil

^[8] Augusta Univ, Dent Coll Georgia, Augusta, GA - USA

^[9] Univ Mogi das Cruzes, Interdisciplinary Ctr Biochem Invest, Mogi Das Cruzes, SP - Brazil

• Tipo de documento: Artigo Científico
• Fonte: ARCHIVES OF ORAL BIOLOGY; v. 117, SEP 2020.
• Citações Web of Science: 0

Resumo

Objective: Cysteine proteases are lysosomal enzymes that, under specific circumstances, may be secreted into the extracellular space and participate in protein turnover. This study investigated the involvement of cathepsin B in the gelatinolytic activity of mature dentin matrices at neutral pH. Design: Human dentin fragments were made into powder and enzymes were extracted using guanidine-HCl/EDTA. Host-derived dentin proteases (cathepsin B, MMP-2 and MMP-9) were identified by immunoblotting, and their activities were evaluated spectrofluorimetrically using fluorogenic substrates. Proteases activities were monitored by measuring the rate of hydrolysis of substrates in the presence/absence of MMP- or cysteine cathepsin inhibitors, at neutral pH (7.4). Mass spectroscopy was used to determine the substrates' cleavage points. Reverse zymography was performed to examine the gelatinolytic activity of cathepsin B. Results: Western-blots of dentin extracts yielded strong bands at 95, 72 and 30 kDa, corresponding respectively to MMP-9, MMP-2 and Cathepsin B. Greater fluorogenic substrates hydrolysis occurred in the absence of MMP and cysteine cathepsin inhibitors than in their presence. Cathepsin B exhibited significant gelatinolytic activity. Conclusions: Together with MMP-2 and MMP-9, cathepsin B also account for the host-derived gelatinolytic activity and matrix turnover of mature dentin at physiological, neutral pH. (AU)

• Processo FAPESP: 16/07996-2 - Síntese e caracterização de um cimento odontológico experimental obtido a partir de extrato de algas marinhas vermelhas
• Beneficiário: Marcela Rocha de Oliveira Carrilho
• Modalidade de apoio: Auxílio à Pesquisa - Regular
• Processo FAPESP: 13/05822-9 - Receptores ativados por proteases (PARs) no complexo dentina-polpa: identificação, modulação e sinalização celular durante o desenvolvimento da doença cárie
• Beneficiário: Fábio Dupart Nascimento
• Modalidade de apoio: Auxílio à Pesquisa - Jovens Pesquisadores
• Processo FAPESP: 11/12226-8 - Avaliação da atividade de proteases na degradação da dentina humana tratada por procedimentos restauradores adesivos
• Beneficiário: Marcela Rocha de Oliveira Carrilho
• Modalidade de apoio: Bolsas no Exterior - Pesquisa