AVALIAÇÃO DE PROTEÍNAS DE CHOQUE TÉRMICO EM CÉLULAS NEOPLÁSICAS DE LARINGE (HEP-2) APÓS TRATAMENTO FOTODINÂMICO
DOI:
https://doi.org/10.18066/revistaunivap.v25i48.2259
Abstract
Proteínas de choque térmico (HSP) são moléculas intracelulares multifuncionais que, eventualmente, podem estar envolvidas na malignização celular. Terapia fotodinâmica (TFD) pode levar à redução do tumor e vasos sanguíneos ao redor. Objetivo foi avaliar ação da TFD sobre as HSPs em células neoplásicas de carcinoma de laringe humana (HEp-2). As células foram irradiadas com LED a 660 nm, 5 J/cm2, 70 mW, por 3 minutos e 20 segundos; incubadas por períodos de 24, 48 e 72 horas; após os períodos de incubação foi realizada a extração de proteínas e corrido gel de poliacrilamida para avaliação das proteínas por Western-Blotting. As células foram imunomarcadas com anticorpos anti-HSP27, anti-HSP70 e anti-HSP90 e analisadas no microscópio confocal. A viabilidade celular foi avaliada pelo teste de cristal violeta. No gel de poliacrilamida foram identificadas bandas próximas a 27 kDa, 70 kDa e 90 kDa. Nas fotomicrografias observou-se redução do número de células após TFD, comprovado por teste de viabilidade (cristal violeta); e intensa marcação de HSPs após TFD, principalmente próximas ao núcleo. Concluiu-se que HSP27, HSP70 e HSP90 são muito produzidas em células HEp-2. TFD foi eficaz devido à redução do número de células e considerada opção viável para o tratamento dessa doença. Embora seja relatada a participação das HSPs 27, 70 e 90 como protetoras das células tumorais, nossos resultados indicam que a TFD ativa tais proteínas para a redução das células tumorais.
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References
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BASSO, F. G., PANSANI, T. N., SOARES, D. G., SCHEFFEL, D. L., BAGNATO, V. S., DE, SOUZA COSTA, C. A. and HEBLING, J. Biomodulation of Inflammatory Cytokines Related to Oral Mucositis by Low‐Level Laser Therapy. Photochem Photobiol, v. 91, n. 4, p. 952-956, 2015. doi:10.1111/php.12445.
BLUM, J. S.; WEARSCH, P. A.; CRESSWELL, P. Pathways of Antigen Processing. Annual Review of Immunology, v. 31, n. 1, p. 443–473, 2013.
BROEKGAARDEN, M. et al. Tumor cell survival pathways activated by photodynamic therapy: a molecular basis for pharmacological inhibition strategies. Cancer and Metastasis Reviews, v. 34, n. 4, p. 643-690, 2015.
BUTTROS, D. A. B. Associação das HSP 60 e 70 com fatores de risco cardiovascular em mulheres na pós-menopausa tratadas de câncer de mama. 2018. Tese (Doutorado) - Universidade Estadual Paulista Júlio de Mesquita Filho, Botucatu – SP, 2018. Disponível em: https://repositorio.unesp.br/handle/11449/154253. Acesso em: 26 fev. 2019.
CALDERWOOD, S. K. Heat shock proteins in breast cancer progression–A suitable case for treatment? International Journal of Hyperthermia, v. 26, n. 7, p. 681- 685, 2010.
CASAS, A. et al. Mechanisms of Resistance to Photodynamic Therapy. Current Medicinal Chemistry, v. 18, n. 16, p. 2486-2515, 2011.
COHEN-SFADY, M. et al. Heat Shock Protein 60, via MyD88 Innate Signaling, Protects B Cells from Apoptosis, Spontaneous and Induced. The Journal of Immunology, v. 183, n. 2, p. 890-896, 2009.
ETMINAN, N. et al. Modulation of migratory activity and invasiveness of human glioma spheroids following 5-aminolevulinic acid–based photodynamic treatment. Journal of Neurosurgery, v. 115, n. 2, p. 281–288, 2011.
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2019-12-06
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Soares, C. P., Uchôas, M. H., Godoi, B. H., & Moraes, C. D. G. de O. (2019). AVALIAÇÃO DE PROTEÍNAS DE CHOQUE TÉRMICO EM CÉLULAS NEOPLÁSICAS DE LARINGE (HEP-2) APÓS TRATAMENTO FOTODINÂMICO. Revista Univap, 25(48), 43–63. https://doi.org/10.18066/revistaunivap.v25i48.2259
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DOI:
https://doi.org/10.18066/revistaunivap.v25i48.2259Abstract
Proteínas de choque térmico (HSP) são moléculas intracelulares multifuncionais que, eventualmente, podem estar envolvidas na malignização celular. Terapia fotodinâmica (TFD) pode levar à redução do tumor e vasos sanguíneos ao redor. Objetivo foi avaliar ação da TFD sobre as HSPs em células neoplásicas de carcinoma de laringe humana (HEp-2). As células foram irradiadas com LED a 660 nm, 5 J/cm2, 70 mW, por 3 minutos e 20 segundos; incubadas por períodos de 24, 48 e 72 horas; após os períodos de incubação foi realizada a extração de proteínas e corrido gel de poliacrilamida para avaliação das proteínas por Western-Blotting. As células foram imunomarcadas com anticorpos anti-HSP27, anti-HSP70 e anti-HSP90 e analisadas no microscópio confocal. A viabilidade celular foi avaliada pelo teste de cristal violeta. No gel de poliacrilamida foram identificadas bandas próximas a 27 kDa, 70 kDa e 90 kDa. Nas fotomicrografias observou-se redução do número de células após TFD, comprovado por teste de viabilidade (cristal violeta); e intensa marcação de HSPs após TFD, principalmente próximas ao núcleo. Concluiu-se que HSP27, HSP70 e HSP90 são muito produzidas em células HEp-2. TFD foi eficaz devido à redução do número de células e considerada opção viável para o tratamento dessa doença. Embora seja relatada a participação das HSPs 27, 70 e 90 como protetoras das células tumorais, nossos resultados indicam que a TFD ativa tais proteínas para a redução das células tumorais.Downloads
References
AGOSTINIS, P. et al. Photodynamic therapy of cancer: An update. CA: A Cancer Journal for Clinicians, v. 61, n. 4, p. 250–281, 2011.
BASSO, F. G., PANSANI, T. N., SOARES, D. G., SCHEFFEL, D. L., BAGNATO, V. S., DE, SOUZA COSTA, C. A. and HEBLING, J. Biomodulation of Inflammatory Cytokines Related to Oral Mucositis by Low‐Level Laser Therapy. Photochem Photobiol, v. 91, n. 4, p. 952-956, 2015. doi:10.1111/php.12445.
BLUM, J. S.; WEARSCH, P. A.; CRESSWELL, P. Pathways of Antigen Processing. Annual Review of Immunology, v. 31, n. 1, p. 443–473, 2013.
BROEKGAARDEN, M. et al. Tumor cell survival pathways activated by photodynamic therapy: a molecular basis for pharmacological inhibition strategies. Cancer and Metastasis Reviews, v. 34, n. 4, p. 643-690, 2015.
BUTTROS, D. A. B. Associação das HSP 60 e 70 com fatores de risco cardiovascular em mulheres na pós-menopausa tratadas de câncer de mama. 2018. Tese (Doutorado) - Universidade Estadual Paulista Júlio de Mesquita Filho, Botucatu – SP, 2018. Disponível em: https://repositorio.unesp.br/handle/11449/154253. Acesso em: 26 fev. 2019.
CALDERWOOD, S. K. Heat shock proteins in breast cancer progression–A suitable case for treatment? International Journal of Hyperthermia, v. 26, n. 7, p. 681- 685, 2010.
CASAS, A. et al. Mechanisms of Resistance to Photodynamic Therapy. Current Medicinal Chemistry, v. 18, n. 16, p. 2486-2515, 2011.
COHEN-SFADY, M. et al. Heat Shock Protein 60, via MyD88 Innate Signaling, Protects B Cells from Apoptosis, Spontaneous and Induced. The Journal of Immunology, v. 183, n. 2, p. 890-896, 2009.
ETMINAN, N. et al. Modulation of migratory activity and invasiveness of human glioma spheroids following 5-aminolevulinic acid–based photodynamic treatment. Journal of Neurosurgery, v. 115, n. 2, p. 281–288, 2011.
EVANS, C. G.; CHANG, L.; GESTWICKI, J. E. Heat Shock Protein 70 (Hsp70) as an Emerging Drug Target. Journal of Medicinal Chemistry, v. 53, n. 12, p. 4585–4602, 2010.
FEOKTISTOVA, M., GESERICK, P. AND LEVERKUS, M. Crystal Violet Assay for Determining Viability of Cultured Cells. Cold Spring Harbor Protocols, n. 4, 2016, pdb.prot087379. https://doi.org/10.1101/pdb.prot087379
GRACEFFA, P. Hsp27-Actin Interaction. Biochemistry Research International, v. 2011, p. 1–7, 2010.
GRANER, M. W. et al. Endoplasmic Reticulum Chaperones and Their Roles in the Immunogenicity of Cancer Vaccines. Frontiers in Oncology, v. 4, 2015.
GUERRERO-ROJAS, R; GUERRERO-FONSECAZ, C. Mecanismos moleculares de las proteínas de choque térmico (HSPs) implicados en el desarrollo neoplásico. Salud Uninorte. v. 34, n. 2, p. 455-474, 2018. Disponível em: http://rcientificas.uninorte.edu.co/index.php/salud/article/view/7160/214421443 429. Acesso em: 26 fev. 2019.
GUO, F. Mechanistic role of heat shock protein 70 in Bcr-Abl-mediated resistance to apoptosis in human acute leukemia cells. Blood, v. 105, n. 3, p. 1246–1255, 2004.
HELBIG, D.; SIMON, J. C.; PAASCH, U. Photodynamic therapy and the role of heat shock protein 70. International Journal of Hyperthermia, v. 27, n. 8, p. 802-810, 2011.
HUANG, L. et al. A tumor-targeted Ganetespib-zinc phthalocyanine conjugate for synergistic chemo-photodynamic therapy. European Journal of Medicinal Chemistry, v. 151, 294-303, 2018.
ISSA, MCA; MANELA-AZULAY, M. Terapia fotodinâmica: revisão da literatura e documentação iconográfica. Anais Brasileiros de Dermatologia, v. 85, n. 4, p. 501-511, 2010.
JEGO, G. et al. Targeting heat shock proteins in cancer. Cancer Letters, v. 332, n. 2, p. 275-285, 2013.
JUHASZ, K. et al. The Complex Function of Hsp70 in Metastatic Cancer. Cancers, v. 6, n. 1, p. 42-66, 2013.
KALMAR, B; GREENSMITH, L. Induction of heat shock proteins for protection against oxidative stress. Advanced Drug Delivery Reviews, v. 61, n. 4, p. 310-318, 2009.
KANEKO, K. et al. Induction of enhanced tumor-specific immunity by Hsp90 targeted photodynamic therapy (Hsp90-PDT) combined with immune checkpoint inhibition. AACR Annual Meeting, Washington, 2017.
KAPOOR, C.; VAIDYA, S. Heat shock protein (HSP) and cancer: an overview. Am. J. Med. Dent. Sci., v. 1, n. 1, p. 31-34, 2013.
KARA, C, SELAMET, H, GÖKMENOĞLU, C, KARA, N. Low level laser therapy induces increased viability and proliferation in isolated cancer cells. Cell Prolif. v. 51, n. 2, e12417, 2018. https://doi.org/10.1111/cpr.12417
KHALIL, A. A. et al. Heat shock proteins in oncology: Diagnostic biomarkers or therapeutic targets? Biochimica et Biophysica Acta (BBA) - Reviews on Cancer, v. 1816, n. 2, p. 89-104, 2011.
KIM, J. et al. Effects of HSP27 downregulation on PDT resistance through PDT-induced autophagy in head and neck cancer cells. Oncology Reports, v. 35, n. 4, p. 2237-2245, 2016.
KIM, J. et al. Down-regulation of heat-shock protein 27-induced resistance to photodynamic therapy in oral cancer cells. Journal of Oral Pathology & Medicine, v. 42, n. 1, p. 9-16, 2012.
KOCAMAN G, BELDÜZ N, ERDOGAN C, OZBEK E, SADIK E, KARA C. The use of surgical Nd:YAG laser in an oral pyogenic granuloma: a case report. J Cosmet Laser Ther. v.16, n. 4, p. 197‐200, 2014.
KOSTENKO, S.; MOENS, U. Heat shock protein 27 phosphorylation: kinases, phosphatases, functions and pathology. Cellular and Molecular Life Sciences, v. 66, n. 20, p. 3289-3307, 2009.
KUMAR, S. et al. Targeting Hsp70: A possible therapy for cancer. Cancer Letters, v. 374, n. 1, p. 156-166, 2016.
LI, Y. et al. Nanotechnology-based photoimmunological therapies for cancer. Cancer Letters, v. 442, p. 429-438, 2019.
LONG, Q. et el. Image-guided photo-therapeutic nanoporphyrin synergized HSP90 inhibitor in patient-derived xenograft bladder cancer model. Nanomedicine: Nanotechnology, Biology and Medicine, v. 14, n. 3, p. 789-799, 2018.
LORENZO, J. G.; VINTRÓ, X. L.; MADRID, M. C. P. Expresión de Heat Shock Protein-90 (HSP-90) como factor predictor de la respuesta a radioterapia en pacientes con tumores de cabeza y cuello. Acta Otorrinolaringológica Española, v. 67, n. 3, p. 130-134, 2016.
LU, X.; KAKKAR, V. The Role of Heat Shock Protein (HSP) in Atherosclerosis: Pathophysiology and Clinical Opportunities. Current Medicinal Chemistry, v. 17, n. 10, p. 957-973, 2010.
MAFTOUM‐COSTA, M., NAVES, K. T., OLIVEIRA, A. L., TEDESCO, A. C., DA, SILVA, N. S. AND PACHECO‐SOARES, C. Mitochondria, endoplasmic reticulum and actin filament behavior after PDT with chloroaluminum phthalocyanine liposomal in HeLa cells. Cell Biology International, v. 32, n. 8, p. 1024-1028. 2008, doi:10.1016/j.cellbi.2008.04.005
MIGLIORATI C., HEWSON I. et al. Systematic review of laser and other light therapy for the management of oral mucositis in cancer patients. Supportive Care in Cancer, v. 21, n. 1, p. 333-341, 2013.
MORAES, C. D. G. O. et al. Genotoxic effects of photodynamic therapy in laryngeal cancer cells – An in vitro study. Experimental Biology and Medicine, v. 244, n. 3, p. 262-271, 2019.
MULLER, C. H. L. Resposta ao choque térmico e da razão HSPA1A extra/intraceluar em leucócitos de indivíduos idosos e de meia idade submetidos a treinamento de força. 2018. Dissertação (Mestrado) - Universidade Federal do Rio Grande do Sul, Porto Alegre, 2018. Disponível em: https://www.lume.ufrgs.br/bi tstream/handle/10183/174954/001063817.pdf?sequence=1&isAllowed=y. Acesso em: 26 fev. 2019.
OLSEN J.G., TEILUM K et al. Behaviour of intrinsically disordered proteins in protein–protein complexes with an emphasis on fuzziness. Cellular and Molecular Life Sciences, v. 74, n. 17, p. 3175-3183, 2017.
PANZARINI, E.; INGUSCIO, V.; DINI, L. Immunogenic Cell Death: Can It Be Exploited in PhotoDynamic Therapy for Cancer? BioMed Research International, v. 2013, p. 1-18, 2013.
RADONS, J. The human HSP70 family of chaperones: where do we stand? Cell Stress and Chaperones, v. 21, n. 3, p. 379-404, 2016.
RAMOS, L. P. Atividade antimicrobiana e citotoxicidade dos extratos glicólicos de Pfaffia paniculata e Juglans regia L. São José dos Campos: [s.n.], 2016.
ROBERTSON CA, EVANS DH, ABRAHAMSE H. Photodynamic therapy (PDT): a short review on cellular mechanisms and cancer research applications for PDT. J Photochem Photobiol B, v. 96, n.1, p. 1-8, 2009.
RODRÍGUEZ, M. E. et al. Heat shock proteins in the context of photodynamic therapy: autophagy, apoptosis and immunogenic cell death. Photochemical & Photobiological Sciences, v. 15, n. 9, p. 1090-1102, 2016.
SCHLECHT, R. et al. Functional Analysis of Hsp70 Inhibitors. PLoS ONE, v. 8, n. 11, 2013.
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