The role of HIF-1 Alpha signaling in the retina under hypoxic conditions

Thu, 11/03/2016 - 13:52

Hypoxia inducible factor 1 (HIF-1) is a protein that plays an essential role in hypoxia, or low levels of cellular oxygen. HIF-1 is a heterodimeric protein that consists of a constitutively expressed beta subunit and oxygen related alpha subunit.  Both subunits have a basic helix-loop-helix domain that leads to dimerization, where HIF-1 alpha carries an oxygen-dependent degradation (ODD) domain.  Outside of the role of HIF-1 in hypoxia adaptation, it is also involved in cancer, inflammation and metabolism.  The HIF-1 alpha subunit is specifically degraded in normal oxygen environments, where it is not active and functioning until a hypoxic environment is sensed.  In some cases, HIF-1 alpha expression can be quite high, leading to angiogenesis and increased blood flow.  The link between HIF-1 alpha activity and retinal pathways has long been established, given that photoreceptors in the retina require a large amount of oxygen to function properly.  The use of a HIF-1 alpha antibody in the research of HIF-1 alpha signaling and behavior in the retina is highlighted below. 

HIF-1 alpha antibody

Western Blot: HIF-1 alpha Antibody (H1alpha67) [NB100-105] - Analysis using the HRP conjugate of NB100-105. Detection of 50ug cobalt chloride induced COS-7 nuclear extracts (NB800-PC26) using NB100-105.

To begin, Paeng et al used a HIF-1 alpha antibody to help elucidate the role of caffeic acid phenethyl ester (CAPE) in vascular endothelial growth factor reduction in the human retinal pigment epithelium under hypoxic conditions.  First, a HIF-1 alpha antibody was used in western blot on ARPE-19 cells that had been treated with CAPE.  This experiment determined that translocation of HIF-1 alpha to the nucleus in hypoxic ARPE-19 cells was nearly two-fold of that in normoxic conditions.  However, with pretreatment of CAPE, the translocation of HIF-1 alpha to the nucleus was greatly prevented.  The HIF-1 alpha antibody was also used in western blot to test the effects of a known number of inhibitory signaling transduction pathways.  Interestingly, AKT had an increase in expression after 2 hours of increased hypoxia, however pre-treatment with CAPE reduced hypoxia-induced AKT phosphorylation.  Overall, it was determined that HIF-1 alpha translocates to the nucleus, binds to the hypoxia response element of VEGF, and leads to ocular angiogenesis.

Next, Park et al used a HIF-1 antibody to show that 3,3’-Diindolylmethane (DIM) inhibits VEGF expression through HIF-1 alpha and NF-κB pathways in the retinal epithelium in hypoxic conditions.  For starters, the HIF-1 alpha antibody was used in western blot analysis of retinal pigment epithelial (RPE) cells to show that HIF-1 alpha translocates to the nucleus under hypoxic conditions, which is the foundational step of hypoxia experiments.  Subsequently, the interaction and binding of HIF-1 alpha and NF-κB was solidified through an electrophoretic mobility shift assay.  A HIF-1 alpha antibody was also used to show that translocation of HIF-1 alpha and NF-κB in this model was halted by introduction to CoCl2 treatment.  Ultimately it was hypothesized that pre-treatment of RPE cells in hypoxic conditions with a ROS inhibitor, caused a down regulation of VEGF through the inhibition of HIF-1 alpha. 

Novus Biologicals offers HIF-1 alpha reagents for your research needs including:

PMID: 25738890
PMID: 25955241
PMID: 18160990

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