Hypoxia causes a large array of adaptive and physiological responses in all cells including cardiac myocytes. In order to elucidate the molecular effects of increased glucose flux on hypoxic cardiac myocytes we focused on the basic helix-loop-helix transcription factor, hypoxia inducible factor 1 alpha (HIF-1α), which is rapidly upregulated in hypoxic cells and elicits a number of responses including augmentation of glucose uptake. Primary cultures of neonatal rat cardiac myocytes as well as embryonic rat heart-derived myogenic H9c2 cells demonstrated a significant upregulation of HIF-1α when subjected to hypoxia of 6-8 h in the absence of glucose. Re-addition of extracellular glucose to the medium resulted in a decrease of HIF-1α levels by almost 50%. This glucose effect was blocked by addition of glycolytic inhibitors. In addition, glucose uptake and glycolysis resulted in substantial decreased levels of p53, which is regulated by HIF-1α. Adenoviral infection of cultures of cardiac myocytes with the facilitative glucose transporter, GLUT1 followed by hypoxia of 24 h also resulted in a significant reduction in the protein expression of HIF-1α compared to control vector-infected cultures. GLUT1 infected cultures also demonstrated fewer apoptotic cells and a reduction in the release of cytochrome c after hypoxia. Inhibition of the ubiquitin-proteasomal pathway by a variety of 26S proteasomal inhibitors increased HIF-1α to similar levels under both normoxic and hypoxic conditions and in the presence or absence of glucose. This result suggested that glucose induces HIF-1α degradation via a proteasomal pathway. This conclusion was substantiated by immuno-precipitation experiments of total cell extracts, which demonstrated an increase of ubiquitinated HIF-1α relative to total HIF-1α in the presence of glucose during hypoxia. Thus, glucose as well as GLUT1 overexpression diminishes hypoxia-induced HIF-1α protein via an ubiquitin-proteasomal pathway in hypoxic cardiac myocytes. This represents a novel feedback mechanism that may play an important role in adaptation of cardiac myocytes to hypoxia and ischemia.
- Cardiac myocytes
ASJC Scopus subject areas
- Molecular Biology
- Cardiology and Cardiovascular Medicine