Dataset Information

Double oxygen-sensing vector system for robust hypoxia/ischemia-regulated gene induction in cardiac muscle in vitro and in vivo.

ABSTRACT: High-fidelity genetically encoded bio-sensors that respond to changes in cellular environmental milieu in disease offer great potential in a range of patho-physiological settings. Here a unique hypoxia-regulated vector-based system with double oxygen-sensing transcriptional elements was developed for rapid and robust hypoxia-regulated gene expression in the heart. Hypoxia-responsive cis elements were used in tandem with a single proline-modified oxygen-dependent degradation (ODD) domain of hypoxia-inducible factor-1alpha to form a double oxygen-sensing vector system (DOSVS). In adult cardiac myocytes in vitro, the DOSVS demonstrated a low background expression not different from baseline control in normoxia, and with 100% efficiency, robust, 1,000-fold induction upon hypoxia. In the heart in vivo, hypoxic and ischemic challenges elicited rapid 700-fold induction in living animals, exceeding that obtained by a high-fidelity constitutive cytomegalovirus (CMV) viral promoter. DOSVS also showed high temporal resolution in the heart in response to cyclical bouts of hypoxia in vivo. We propose that DOSVS will be valuable for a range of applications, including bio-sensing and therapeutic gene expression in the heart and other organ systems that are confronted by chronic or episodic hypoxic/ischemic stresses in vivo.

SUBMITTER: Fomicheva EV

PROVIDER: S-EPMC2716210 | biostudies-literature | 2008 Sep

REPOSITORIES: biostudies-literature

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Double oxygen-sensing vector system for robust hypoxia/ischemia-regulated gene induction in cardiac muscle in vitro and in vivo.

Fomicheva Ekaterina V EV Turner Immanuel I II Edwards Terri G TG Hoff Janet J Arden Eric E D'Alecy Louis G LG Metzger Joseph M JM

Molecular therapy : the journal of the American Society of Gene Therapy 20080624 9

High-fidelity genetically encoded bio-sensors that respond to changes in cellular environmental milieu in disease offer great potential in a range of patho-physiological settings. Here a unique hypoxia-regulated vector-based system with double oxygen-sensing transcriptional elements was developed for rapid and robust hypoxia-regulated gene expression in the heart. Hypoxia-responsive cis elements were used in tandem with a single proline-modified oxygen-dependent degradation (ODD) domain of hypox ...[more]

PMID: 18578010

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Project description:As the ultimate electron acceptor in oxidative phosphorylation, oxygen plays a critical role in metabolism. When oxygen levels drop, heterodimeric hypoxia-inducible factor (Hif) transcription factors become active and facilitate adaptation to hypoxia. Hif regulation by oxygen requires the protein von Hippel-Lindau (pVhl) and pVhl disruption results in constitutive Hif activation. The liver is a critical organ for metabolic homeostasis, and Vhl inactivation in hepatocytes results in a Hif-dependent shortening in life span. While albumin-Cre;Vhl(F/F) mice develop hepatic steatosis and impaired fatty acid oxidation, the variable penetrance and unpredictable life expectancy has made the cause of death elusive. Using a system in which Vhl is acutely disrupted and a combination of ex vivo liver perfusion studies and in vivo oxygen measurements, we demonstrate that Vhl is essential for mitochondrial respiration in vivo. Adenovirus-Cre mediated acute Vhl disruption in the liver caused death within days. Deprived of pVhl, livers accumulated tryglicerides and circulating ketone and glucose levels dropped. The phenotype was reminiscent of inborn defects in fatty acid oxidation and of fasted PPAR?-deficient mice and while death was unaffected by pharmacologic PPAR? activation, it was delayed by glucose administration. Ex vivo liver perfusion analyses and acylcarnitine profiles showed mitochondrial impairment and a profound inhibition of liver ketone and glucose production. By contrast, other mitochondrial functions, such as ureagenesis, were unaffected. Oxygen consumption studies revealed a marked suppression of mitochondrial respiration, which, as determined by magnetic resonance oximetry in live mice, was accompanied by a corresponding increase in liver pO(2). Importantly, simultaneous inactivation of Hif-1? suppressed liver steatosis and rescued the mice from death. These data demonstrate that constitutive Hif activation in mice is sufficient to suppress mitochondrial respiration in vivo and that no other pathway exists in the liver that can allow oxygen utilization when Hif is active precluding thereby metabolic collapse.

Dataset Information

Double oxygen-sensing vector system for robust hypoxia/ischemia-regulated gene induction in cardiac muscle in vitro and in vivo.

Publications

Double oxygen-sensing vector system for robust hypoxia/ischemia-regulated gene induction in cardiac muscle in vitro and in vivo.

Similar Datasets

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