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Dynamic imaging of autophagy-lysosomal pathway and autophagy function following pulmonary hypoxia/reoxygenation in vitro / 华中科技大学学报(医学)(英德文版)
Journal of Huazhong University of Science and Technology (Medical Sciences) ; (6): 302-308, 2015.
Artigo em Inglês | WPRIM | ID: wpr-331069
ABSTRACT
Alterations of the autophagy-lysosomal pathway (ALP) and autophagy have been involved in lung ischemia-reperfusion (I/R) injury. However, dynamic imaging of ALP function under lung I/R injury particularly is not fully understood. Here we depicted the live-cell fluorescence imaging of autophagosome to monitor ALP activation and autophagy function. The pAsRed2-N1-LC3 vectors were transfected into CRL-2192 NR8383 (an alveolar macrophage cell line) and CCL149 (an alveolar epithelial cell line) successfully. 0-h, 2-h, 4-h, and 6-h hypoxia/0-h, 2-h, 4-h, and 6-h reoxygenation were then induced with an ALP inhibitor (3-MA) or activator (rapamycin) in the culture of transfected cells separately. ALP activation was conformed by up-regulating AMPK and beclin1 expression. Apoptosis was not obvious in 2-h hypoxia/2-h reoxygenation. pAsRed2-N1-LC3 CCL149 and pAsRed2-N1-LC3 NR8383 cells revealed gradually enhanced AsRed2 from 2-h to 6-h hypoxia/reoxygenation. AsRed2 varied sensitively to 3-MA and rapamycin interventions during 2-h hypoxia/reoxygenation. Our data provides a simple method of autophagosome imaging to monitor ALP activation and autophagy function in lung I/R injury.
Assuntos
Texto completo: DisponíveL Índice: WPRIM (Pacífico Ocidental) Assunto principal: Oxigenoterapia / Fisiologia / Autofagia / Técnicas In Vitro / Sequência de Bases / Primers do DNA / Reação em Cadeia da Polimerase em Tempo Real / Pulmão / Lisossomos / Hipóxia Limite: Animais Idioma: Inglês Revista: Journal of Huazhong University of Science and Technology (Medical Sciences) Ano de publicação: 2015 Tipo de documento: Artigo

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Texto completo: DisponíveL Índice: WPRIM (Pacífico Ocidental) Assunto principal: Oxigenoterapia / Fisiologia / Autofagia / Técnicas In Vitro / Sequência de Bases / Primers do DNA / Reação em Cadeia da Polimerase em Tempo Real / Pulmão / Lisossomos / Hipóxia Limite: Animais Idioma: Inglês Revista: Journal of Huazhong University of Science and Technology (Medical Sciences) Ano de publicação: 2015 Tipo de documento: Artigo