Histomorphometric analysis of the lung of Swiss mice treated with a fibrinolytic protease

Fibrinolytic enzymes are considered promising alternative in the treatment of cardiovascular diseases by preventing fibrin clots. A protease from Mucor subtilissimus UCP 1262 was obtained by solid state fermentation and purified by ion exchange chromatography. The purified extract was administered at an acute dose of 2000 mg/mL to evaluate its toxic effects to the lungs of mice. After 14 days of treatment, a histomorphometric study was performed by the type 1 and 2 pneumocyte count and the evaluation of the lung area. As result, the experimental group showed a significant decrease of type 2 pneumocyte and although a decrease in the alveolar area was observed in relation to the control group, no significant pulmonary toxicity, emphysema, and fibrosis characteristics were detected. The in vitro tests suggest possible clinical applications for the enzyme.

Key words
Fibrinolytic protease; thrombosis; Mucor; pneumocyte; thrombolytic agent

Authorship

MARLLYN M. DA SILVA

Universidade Federal Rural de Pernambuco, Laboratório de Tecnologia de Bioativos, Departamento de Morfologia e Fisiologia Animal, Rua Dom Manuel de Medeiros, s/n, Dois Irmãos, 52171-900 Recife, PE, BrazilUniversidade Federal Rural de PernambucoBrazilRecife, PE, BrazilUniversidade Federal Rural de Pernambuco, Laboratório de Tecnologia de Bioativos, Departamento de Morfologia e Fisiologia Animal, Rua Dom Manuel de Medeiros, s/n, Dois Irmãos, 52171-900 Recife, PE, Brazil

http://orcid.org/0000-0002-1768-9831

MARIA A.C. DE LIRA

Universidade Federal de Pernambuco, Laboratório de Síntese e Isolamento Molecular, Centro Acadêmico de Vitória de Santo Antão, Rua do Alto do Reservatório, Alto José Leal, 55608-680 Vitória de Santo Antão, PE, BrazilUniversidade Federal de PernambucoBrazilVitória de Santo Antão, PE, BrazilUniversidade Federal de Pernambuco, Laboratório de Síntese e Isolamento Molecular, Centro Acadêmico de Vitória de Santo Antão, Rua do Alto do Reservatório, Alto José Leal, 55608-680 Vitória de Santo Antão, PE, Brazil

http://orcid.org/0000-0002-2662-1003

TAMIRIS A. ROCHA

Universidade Federal de Pernambuco, Nanotecnologia, biotecnologia e cultura de células (Nanobiocel), Centro Acadêmico de Vitória de Santo Antão, Rua do Alto do Reservatório, Alto José Leal, 55608-680 Vitória de Santo Antão, PE, BrazilUniversidade Federal de PernambucoBrazilVitória de Santo Antão, PE, BrazilUniversidade Federal de Pernambuco, Nanotecnologia, biotecnologia e cultura de células (Nanobiocel), Centro Acadêmico de Vitória de Santo Antão, Rua do Alto do Reservatório, Alto José Leal, 55608-680 Vitória de Santo Antão, PE, Brazil

http://orcid.org/0000-0001-5961-6377

DANIELLE F. DE MOURA

Universidade Federal de Pernambuco, Laboratório de Parasitologia, Centro Acadêmico de Vitória de Santo Antão, Rua do Alto do Reservatório, Alto José Leal, 55608-680 Vitória de Santo Antão, PE, BrazilUniversidade Federal de PernambucoBrazilVitória de Santo Antão, PE, BrazilUniversidade Federal de Pernambuco, Laboratório de Parasitologia, Centro Acadêmico de Vitória de Santo Antão, Rua do Alto do Reservatório, Alto José Leal, 55608-680 Vitória de Santo Antão, PE, Brazil

http://orcid.org/0000-0002-8258-0541

FRANCISCO C.A.A. JÚNIOR

Universidade Federal de Pernambuco, Laboratório de Biotecnologia e Fármacos, Centro Acadêmico de Vitória de Santo Antão, Rua do Alto do Reservatório, Alto José Leal, 55608-680 Vitória de Santo Antão, PE, BrazilUniversidade Federal de PernambucoBrazilVitória de Santo Antão, PE, BrazilUniversidade Federal de Pernambuco, Laboratório de Biotecnologia e Fármacos, Centro Acadêmico de Vitória de Santo Antão, Rua do Alto do Reservatório, Alto José Leal, 55608-680 Vitória de Santo Antão, PE, Brazil

http://orcid.org/0000-0001-8676-4826

ANA V.S. FERREIRA

http://orcid.org/0000-0002-9400-4090

LORENZO PASTRANA

International Iberian Nanotechnology Laboratory, Av. Mestre José Veiga, 4715-330 Braga, PortugalInternational Iberian Nanotechnology LaboratoryPortugalAv. Mestre José Veiga, Braga, PortugalInternational Iberian Nanotechnology Laboratory, Av. Mestre José Veiga, 4715-330 Braga, Portugal

http://orcid.org/0000-0002-6637-3462

WENDELL W.C. ALBUQUERQUE

Justus Liebig University, Department of Food Chemistry and Food Biotechnology, 35390, Ludwigstraße 23, Giessen, Germany.Justus Liebig UniversityGermanyLudwigstraße 23, Giessen, GermanyJustus Liebig University, Department of Food Chemistry and Food Biotechnology, 35390, Ludwigstraße 23, Giessen, Germany.

http://orcid.org/0000-0001-7658-491X

ROMERO M.P.B. COSTA

Universidade de Pernambuco, Laboratório de Avanços em Biotecnologia e Proteína (LABIOPROT), Instituto de Ciências Biológicas, Rua Arnóbio Marquês, 310, Santo Amaro, 50100-130 Recife, PE, BrazilUniversidade de PernambucoBrazilRecife, PE, BrazilUniversidade de Pernambuco, Laboratório de Avanços em Biotecnologia e Proteína (LABIOPROT), Instituto de Ciências Biológicas, Rua Arnóbio Marquês, 310, Santo Amaro, 50100-130 Recife, PE, Brazil

http://orcid.org/0000-0001-7045-2975

THIAGO P. NASCIMENTO

Universidade Federal do Piauí, Campus Professora Cinobelina Elvas, BR 135, Km 3, Planalto Horizonte, 64900-000 Bom Jesus, PI, BrazilUniversidade Federal do PiauíBrazilBom Jesus, PI, BrazilUniversidade Federal do Piauí, Campus Professora Cinobelina Elvas, BR 135, Km 3, Planalto Horizonte, 64900-000 Bom Jesus, PI, Brazil

http://orcid.org/0000-0003-3480-6734

ANA L.F. PORTO

http://orcid.org/0000-0001-5561-5158

Correspondence to: Thiago Pajeú Nascimento E-mail: thiago_pajeu@hotmail.com

AUTHOR CONTRIBUTIONS

All authors contributed to the development of the manuscript: Marllyn Marques da Silva– Obtaining fibrinolytic protease, Histomorphometric analysis and writing ; Maria Aparecida da Conceição de Lira - Lung evaluation of acute exposure to fibrinolytic enzyme; Tamiris Alves Rocha - Lung evaluation of acute exposure to fibrinolytic enzyme; Danielle Feijó de Moura - Histomorphometric analysis; Francisco Carlos Amanajás de Aguiar Júnior - Histomorphometric analysis; Ana Vitória da Silva Ferreira - Histomorphometric analysis; Lorenzo Pastrana - help with writing in english; Wendell Wagner Campos Albuquerque - help with writing in english; Romero Marcos Pedrosa Brandão Costa - Obtaining fibrinolytic protease; Thiago Pajeú Nascimento- Analysis of results and assistance in writing them, Ana Lúcia Figueiredo Porto – Analysis of results and assistance in writing them.

SCIMAGO INSTITUTIONS RANKINGS

Figures | Tables

Figures (1)
Tables (1)

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Figure 1
Photomicrographs of lung tissue sections of mice in the different groups studied (H&E). a and c= Preserved lung parenchyma (circle); b and d= Preserved alveolar sac (star), Pneumocyte type 1 (double arrow), Pneumocyte type 2 (arrow). HE staining with magnification of 10X (a and c) and 40X (b and d). Scale bar = 1 μm.

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Table I
Number of type and type 2 pneumocytes in the lungs of mice treated with the fibrinolytic enzyme produced by Mucor subtilissimus UCP 1262 (2000mg/mL). The results are expressed as mean ± SD. Statistical differences were determined by the Mann-Whitney U-test. CN: negative control; ENZ: fibrinolytic enzyme produced by Mucor subtilissimus. *P<0.05 vs Control.

Figure 1 Photomicrographs of lung tissue sections of mice in the different groups studied (H&E). a and c= Preserved lung parenchyma (circle); b and d= Preserved alveolar sac (star), Pneumocyte type 1 (double arrow), Pneumocyte type 2 (arrow). HE staining with magnification of 10X (a and c) and 40X (b and d). Scale bar = 1 μm.

Table I Number of type and type 2 pneumocytes in the lungs of mice treated with the fibrinolytic enzyme produced by Mucor subtilissimus UCP 1262 (2000mg/mL). The results are expressed as mean ± SD. Statistical differences were determined by the Mann-Whitney U-test. CN: negative control; ENZ: fibrinolytic enzyme produced by Mucor subtilissimus. *P<0.05 vs Control.

	Pneumocyte type 1 (n of cells/ µm²)	Pneumocyte type 2 (n of cells/ µm²)	Alveolar Area (n of cells/µm²)
CN	20,38 ± 6,79	29,04 ± 9,58	580,30 ± 230,41
ENZ	17,74 ± 8,44	23,25 ± 11,55*	492,40 ± 242,72 *

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Histomorphometric analysis of the lung of Swiss mice treated with a fibrinolytic protease

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[1] Correspondence to: Thiago Pajeú Nascimento E-mail: thiago_pajeu@hotmail.com