Abstract
Article 1:
Cardioprotection by hypoxia-inducible factor 1 alpha transfection in skeletal muscle is dependent on haem oxygenase activity in mice.
Aims: The present study investigates whether the cardioprotection achieved by gene delivery of hypoxiainducible factor-1a (HIF-1a) depends on the downstream factor haem oxygenase (HMOX)-1.
Methods and results: Immortalized cardiomyocytes (HL-1 cells) were transfected with HIF-1a or HMOX-1 and injured with hydrogen peroxide (H2O2), and death was evaluated by trypan blue staining. Quadriceps muscles of mice were treated with DNA for HIF-1a and HMOX-1, or sham-treated and electroporated, and 3 days later, hearts were isolated and subjected to global ischaemia and reperfusion. Some HIF-1a- and sham-treated mice received the HMOX blocker zinc deuteroporphyrin 2,4-bis-glycol (ZnBG) (n ¼ 6–8 in each group). HL-1 cells were stimulated with bilirubin or the carbon monoxide donor CORM-2 before injury with H2O2. HL-1 cells which were transfected with HIF-1a or HMOX-1 had an increased survival to H2O2-induced injury compared with empty vector (n ¼ 10–12 per group; P , 0.01 for both). When HMOX-1-luciferase reporter mice were treated with HIF-1a in the quadriceps muscle, increased luciferase activity was found locally, but nowhere else. Mice pre-treated with HIF-1a or HMOX-1 had a reduced infarct size, improved post-ischaemic function, and increased serum bilirubin (P , 0.05). ZnBG inhibited all these effects afforded by HIF-1a. Stimulation of HL-1 cells with bilirubin and CORM-2 reduced cell death evoked by H2O2 (P , 0.05 for both, n ¼ 11–15 in each group).
Conclusion HIF-1a and HMOX-1 provided protection against H2O2-induced damage in HL-1 cells. Remote gene delivery of HIF-1a afforded cardioprotective effects. These were dependent on HMOX activity, as an HMOX blocker abolished the effects, and they were mimicked by pre-treatment with HMOX-1. Downstream to HMOX-1, bilirubin as well as carbon monoxide may be organ effectors.
Article2:
Gene therapy with hypoxia-inducible factor 1 alpha in skeletal muscle is cardioprotective in vivo.
Aims: Gene therapy of a peripheral organ to protect the heart is clinically attractive. The transcription factor hypoxia-inducible factor 1 alpha (HIF-1α) transactivates cardioprotective genes. We investigated if remote delivery of DNA encoding for HIF-1α is protective against myocardial ischemia–reperfusion injury in vivo.
Main methods: DNA encoding for human HIF-1α was delivered to quadriceps muscles of mice. One week later myocardial infarction was induced and four weeks later its size was measured. Echocardiography and in vivo pressure–volume analysis was performed. Coronary vascularization was evaluated through plastic casting. HL-1 cells, transfected with either HIF-1α or HMOX-1 or administered bilirubin or the carbon monoxide (CO) donor CORM-2, were subjected to lipopolysacharide (LPS)-induced cell death to compare the efficacy of treatments.
Key findings: After four weeks of reperfusion post infarction, animals pretreated with HIF-1α showed reduced infarct size and left ventricular remodeling (pb0.05, respectively). Fractional shortening was preserved in mice pretreated with HIF-1α (pb0.05). Invasive hemodynamic parameters indicated preserved left ventricular function after HIF-1α (pb0.05), which also induced coronary vascularization (pb0.05). HIF-1α downstream target heme oxygenase 1 (HMOX-1) was upregulated in skeletal muscle, while serum bilirubin was increased. Transfection of HL-1 cells with HIF-1α or HMOX-1 and administration of bilirubin or CORM-2 comparably salvaged cells from lipopolysacharide (LPS)-induced cell death (all pb0.05).
Significance: HIF-1α gene delivery to skeletal muscle preceding myocardial ischemia reduced infarct size and postischemic remodeling accompanied by an improved cardiac function and vascularization. Similar to HIF-1α, HMOX-1, bilirubin and CO were protective against LPS-induced injury. This observation may have clinical potential. cardioprotective in vivo