Heme biosynthesis modulation via δ-aminolevulinic acid administration attenuates chronic hypoxia-induced pulmonary hypertension

Abstract

Alhawaj R, Patel D, Kelly MR, Sun D, Wolin MS. Heme biosynthesis modulation via ␦-aminolevulinic acid administration attenuates chronic hypoxia-induced pulmonary hypertension. Am J Physiol Lung Cell Mol Physiol 308: L719 -L728, 2015. First published February 6, 2015 doi:10.1152/ajplung.00155.2014.-This study examines how heme biosynthesis modulation with ␦-aminolevulinic acid (ALA) potentially functions to prevent 21-day hypoxia (10% oxygen)-induced pulmonary hypertension in mice and the effects of 24-h organoid culture with bovine pulmonary arteries (BPA) with the hypoxia and pulmonary hypertension mediator endothelin-1 (ET-1), with a focus on changes in superoxide and regulation of micro-RNA 204 (miR204) expression by src kinase phosphorylation of signal transducer and activator of transcription-3 (STAT3). The treatment of mice with ALA attenuated pulmonary hypertension (assessed through echo Doppler flow of the pulmonary valve, and direct measurements of right ventricular systolic pressure and right ventricular hypertrophy), increases in pulmonary arterial superoxide (detected by lucigenin), and decreases in lung miR204 and mitochondrial superoxide dismutase (SOD2) expression. ALA treatment of BPA attenuated ET-1-induced increases in mitochondrial superoxide (detected by MitoSox), STAT3 phosphorylation, and decreases in miR204 and SOD2 expression. Because ALA increases BPA protoporphyrin IX (a stimulator of guanylate cyclase) and cGMP-mediated protein kinase G (PKG) activity, the effects of the PKG activator 8-bromo-cGMP were examined and found to also attenuate the ET-1-induced increase in superoxide. ET-1 increased superoxide production and the detection of protoporphyrin IX fluorescence, suggesting oxidant conditions might impair heme biosynthesis by ferrochelatase. However, chronic hypoxia actually increased ferrochelatase activity in mouse pulmonary arteries. Thus, a reversal of factors increasing mitochondrial superoxide and oxidant effects that potentially influence remodeling signaling related to miR204 expression and perhaps iron availability needed for the biosynthesis of heme by the ferrochelatase reaction could be factors in the beneficial actions of ALA in pulmonary hypertension. endothelin; ferrochelatase; guanylate cyclase; micro-RNA 204; superoxide CHRONIC HYPOXIA IS AN IMPORTANT factor in the development of pulmonary hypertension in diseases such as chronic obstructive pulmonary disease (COPD), sleep apnea, and mountain sickness (10,20). It is thought to promote pulmonary hypertension development through persistent pulmonary vasoconstriction and vascular remodeling (31). There is substantial evidence for increases in the generation of reactive oxygen species from sources including Nox oxidases and mitochondria having important roles in the development and/or progression of pulmonary hypertension caused by chronic hypoxia and other stimuli of this disease process (7,15,18,27,30). Pulmonary arterial generation of nitric oxide (NO) is normally thought to oppose this process of vasoconstriction ...

Potential role of mitochondrial superoxide decreasing ferrochelatase and heme in coronary artery soluble guanylate cyclase depletion by angiotensin II

“…For example, mitochondrial superoxide scavengers, which function in a manner similar to Mito-TEMPOL, have the potential for being effective in attenuating the disruption of heme biosynthesis, and Mito-TEMPO has been reported to attenuate vascular dysfunction in the ANG II infusion model (6,17). ALA has already been demonstrated to prevent hypoxia-induced pulmonary hypertension in mice (1,24). Although ALA was not protective in a mouse renal ischemia-reperfusion injury model, when used in combination with ferrous iron, it was protective as a result of generating heme in amounts that appeared to both induce and support the beneficial actions of heme oxygenase (8).…”

“…␤-Actin antibody was purchased from Sigma Chemicals. 1H- [1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ) and spermine-NONOate were purchased from Cayman Chemical (Ann Arbor, MI). Mito-TEMPOL was obtained from Abcam.…”

“…At the beginning of the experiment, a standard curve was generated by injecting increasing concentrations of Mito-2-hydroxyethidium or 2-hydroxyethidium in the column. After 24 h of organoid culture, BCA rings were incubated with either 5 M MitoSox or DHE for 1 h in the dark (1,9) under conditions described in RESULTS to measure mitochondrial and extramitochondrial superoxide, respectively. They were washed several times with Krebs solution buffered with 10 mM HEPES-NaOH (pH 7.4) and then flash-frozen with liquid nitrogen.…”

Redox Mechanisms Influencing cGMP Signaling in Pulmonary Vascular Physiology and Pathophysiology

“…Thus, ET-1 is an important factor generated under conditions leading to PH that contributes to the progression of this disease process. Our studies have found that ET-1 promotes in pulmonary arteries a loss of mitochondrial superoxide dismutase (SOD) expression associated with increased mitochondrial superoxide, and activation other signaling associated with pulmonary arterial smooth muscle remodeling that appear to be prevented by using δ-aminolevulinic acid (ALA) to generate protoporphyrin ix (PpIX), an activator of sGC (10). Treatment of mice with ALA prevented the development of hypoxia-induced PH in mice associated with actions similar to its effects on the actions of ET-1 observed in isolated pulmonary arteries.…”

“…A major interest of our lab has been elucidating aspects of multiple additional mechanisms through which redox can control sGC and cGMP signaling in PA (6–8). Some of these mechanisms seem to participate in pulmonary artery hypoxic pulmonary vasoconstriction (HPV) (6) and changes that occur in pulmonary hypertension (PH) (9, 10). There is now substantial evidence for a loss of endothelium-derived nitric oxide (EDNO) (11) and perhaps its ability to stimulate sGC (12, 13) in various forms of PH.…”