TITLE:
Quantification of the Expression of HIF-1alpha by Real-Time PCR in Rat Hepatocytes Cultures Invaded by Shigella flexneri under Normoxic and Hypoxic Conditions
AUTHORS:
Camila Bárbara Cantalupo Lima, Sânia Alves dos Santos, Dahir Ramos de Andrade Júnior
KEYWORDS:
Shigella flexneri, Hepatocytes, Cell Hypoxia, Hypoxia-Inducible Factor 1α, TNF-Alpha, Apoptosis
JOURNAL NAME:
Advances in Microbiology,
Vol.5 No.7,
July
8,
2015
ABSTRACT: Aim: Shigella flexneri (S. flexneri) is a gram-negative
enterobacterium responsible for severe intestinal end systemic infection in
humans. The bacteria can reach the liver due to degeneration of the colonic
epithelium. Hypoxia is present in many human diseases and can induce the
expression of the transcription factor HIF-1alpha that may have a cell protective
role. The influence of hypoxia and HIF-1alpha on bacterial infection, studied
in this work, is unclear. Hypoxia inducible factor-1alpha (HIF-1alpha) is a
transcription factor that acts as a master regulator of gene expression induced
by hypoxia. Methods: We compared the ability of S. flexneri to invade rat hepatocytes in primary culture both in
normoxic and hypoxic conditions. We evaluated TNF-alpha released by
hepatocytes, apoptosis rate and HIF-1alpha expression by confocal microscopy as
well as real time PCR technique. Results: We showed that S. flexneri invaded less hepatocytes previously submitted to 24 h
hypoxia (6.5% O2) than those cultivated in normoxia (21% O2). S. flexneri also induced HIF-1α expression
in hepatocytes, TNF-α secretion and
apoptosis. Conclusion: a) Hypoxia alone was not a stimulus to TNF-α secretion, but induced cell apoptosis
and HIF-1α expression; b) S. flexneri was able to invade
rat hepatocytes and hypoxia apparently influenced significantly bacterial cell
invasiveness; c) HIF-1α was expressed
in hypoxic conditions, and it was also stimulated by S. flexneri.