Abstract
BackgroundOur previous research found that structural changes of the microtubule network influence glycolysis in cardiomyocytes by regulating the hypoxia-inducible factor (HIF)-1α during the early stages of hypoxia. However, little is known about the underlying regulatory mechanism of the changes of HIF-1α caused by microtubule network alternation. The von Hippel-Lindau tumor suppressor protein (pVHL), as a ubiquitin ligase, is best understood as a negative regulator of HIF-1α.Methodology/Principal FindingsIn primary rat cardiomyocytes and H9c2 cardiac cells, microtubule-stabilization was achieved by pretreating with paclitaxel or transfection of microtubule-associated protein 4 (MAP4) overexpression plasmids and microtubule–depolymerization was achieved by pretreating with colchicine or transfection of MAP4 siRNA before hypoxia treatment. Recombinant adenovirus vectors for overexpressing pVHL or silencing of pVHL expression were constructed and transfected in primary rat cardiomyocytes and H9c2 cells. With different microtubule-stabilizing and -depolymerizing treaments, we demonstrated that the protein levels of HIF-1α were down-regulated through overexpression of pVHL and were up-regulated through knockdown of pVHL in hypoxic cardiomyocytes. Importantly, microtubular structure breakdown activated p38/MAPK pathway, accompanied with the upregulation of pVHL. In coincidence, we found that SB203580, a p38/MAPK inhibitor decreased pVHL while MKK6 (Glu) overexpression increased pVHL in the microtubule network altered-hypoxic cardiomyocytes and H9c2 cells.Conclusions/SignificanceThis study suggests that pVHL plays an important role in the regulation of HIF-1α caused by the changes of microtubular structure and the p38/MAPK pathway participates in the process of pVHL change following microtubule network alteration in hypoxic cardiomyocytes.
Highlights
Hypoxia is a common pathophysiological process in many human diseases
Conclusions/Significance: This study suggests that pVHL plays an important role in the regulation of hypoxia-inducible factor (HIF)-1a caused by the changes of microtubular structure and the p38/Mitogen-activated protein kinase (MAPK) pathway participates in the process of pVHL change following microtubule network alteration in hypoxic cardiomyocytes
The results showed that the HIF-1a protein levels were low under normoxic conditions, and none of the treatments with microtubule-interfering agents produced any change in the HIF-1a protein levels (Figure S1)
Summary
Hypoxia is a common pathophysiological process in many human diseases. Despite being a frequent process in the human system, the hypoxic state is implicated in the onset and progression of many life-threatening diseases. Myocardial hypoxia is a common clinical finding in patients with coronary artery disease, hypertensive heart disease and cardiomyopathy [1,2] This particular type of tissue damage is present in patients with severe burns [3,4]. At the cellular level hypoxia or ischemia elicits cytoskeletal damage, including microtubule network alteration [5,6] This alteration in the sub-cellular architecture in turn influences glycolysis in hypoxic cardiomyocytes (CMs), a process mediated by hypoxia-inducible factor (HIF)-1a [7], which is itself regulated by O2 tension and the microtubule network [7,8,9]. Our previous research found that structural changes of the microtubule network influence glycolysis in cardiomyocytes by regulating the hypoxia-inducible factor (HIF)-1a during the early stages of hypoxia. The von Hippel-Lindau tumor suppressor protein (pVHL), as a ubiquitin ligase, is best understood as a negative regulator of HIF-1a
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