Optimal oxygenation and role of free radicals in PPHN

Abstract

Effective ventilation of the lungs is essential in mediating pulmonary vasodilation at birth to allow effective gas exchange and an increase in systemic oxygenation. Unsuccessful transition prevents the increase in pulmonary blood flow after birth resulting in hypoxemia and persistent pulmonary hypertension of the newborn (PPHN). Management of neonates with PPHN includes ventilation of the lungs with supplemental oxygen to correct hypoxemia. Optimal oxygenation should meet oxygen demand to the tissues and avoid hypoxic pulmonary vasoconstriction (HPV) while preventing oxidative stress. The optimal target for oxygenation in PPHN is not known. Animal models have demonstrated that PaO₂<45 mmHg exacerbates HPV. However, there are no practical methods of assessing oxygen levels associated with oxidant stress. Oxidant stress can be due to free radical generation from underlying lung disease or from free radicals generated by supplemental oxygen. Free radicals act on the nitric oxide pathway reducing cGMP and promoting pulmonary vasoconstriction. Antioxidant therapy improves systemic oxygenation in an animal model of PPHN but there are no clinical trials to support such therapy. Targeting preductal SpO₂ between 90 and 97% and PaO₂ at 50-80 mmHg appears prudent in PPHN but clinical trials to support this practice are lacking. Preterm infants with PPHN present unique challenges due to lack of antioxidant defenses and functional and structural immaturity of the lungs. This review highlights the need for additional studies to mitigate the impact of oxidative stress in the lung and pulmonary vasculature in PPHN.

Figure 1.. Free radicals - reactive oxygen (ROS) and nitrogen species (RNS) in persistent pulmonary hypertension of the newborn.

Reactive oxygen species such as superoxide anions (O₂^.−) can be produced by the electron transport chain (ETC) in the mitochondria, due to exposure to hyperoxia or from enzymes such as uncoupled nitric oxide synthase (NOS), NADPH oxidase, and xanthine oxidase or the Fenton reaction. Nitric oxide (NO) is a free radical and avidly binds to superoxide anion to form peroxynitrite (OONO^.−) at the rate of 6.7/M/s. This rate is considerably faster than the rate of dismutation of superoxide anions by superoxide dismutase (SOD) to form hydrogen peroxide (H₂O₂). Hydrogen peroxide can diffuse across membranes through aquaporins. Hydrogen peroxide is broken down by catalase and glutathione peroxidase (GPx1) to water. NO and oxygen are vasodilators but peroxynitrite is a potent vasoconstrictor. ROS (superoxide anions and hydrogen peroxide) stimulate phosphodiesterase 5 (PDE5) enzyme and breakdown cGMP limiting vasodilator effect of NO. Copyright Satyan Lakshminrusimha. BH4, tetrahydrobiopterin; SOD1 – superoxide dismutase; SOD3, extracellular superoxide dismutase; eNOS, endothelial nitric oxide synthase; SOD2 or MnSOD, manganese superoxide dismutase; sGC, soluble guanylate cyclase; cGMP – cyclic guanosine monophosphate;

Figure 2.. Oxygen tension in different sites of pulmonary circulation and pulmonary vascular resistance (PVR).

The precise site of hypoxic pulmonary vasoconstriction and the sensing mechanisms are not clear but is thought by most to be the precapillary pulmonary arteriole in the lung. The pulmonary arterial smooth muscle cells (PASMC) are exposed to lung tissue PO₂, alveolar PAO₂, and pulmonary arterial (mixed venous) PO₂. It is thought that the rapid diffusion from alveolar PAO₂ is the predominant determinant of oxygen tension in PASMC. In infants with persistent pulmonary hypertension of the newborn (PPHN), PAO₂ can be approximately calculated using preductal PaO₂ values. The presence of a right-to-left shunt at the atrial level or ductal level can reduce PaO₂ levels in PPHN. Heterogeneous lung disease can also interfere with the relationship between alveolar PAO₂ and PVR. Copyright Satyan Lakshminrusimha. Modified from Hemodynamics and Cardiology: Neonatology Questions and Controversies 3^rd Edition.

Figure 3.. Optimal targets for oxygenation in the management of acute PPHN in term infants.

Based on preclinical data, we recommend a preductal PaO₂ of 50 to 80 mmHg in infants with PPHN. The corresponding SpO₂ targets are approximately 90 to 97%. Oxygen targets below this range are associated with hypoxic pulmonary vasoconstriction. Targets above this range are associated with poor response to inhaled nitric oxide and higher incidence of HIE following perinatal depression.

Figure 4.. Cellular and biochemical changes in PPHN secondary to oxidative and nitrosative stress.

Pulmonary arteries from human neonates and animal models of PPHN demonstrate thickening of the muscular layer and adventitia. The normal pulmonary arterial endothelium produces nitric oxide (NO) from phosphorylated endothelial nitric oxide synthase (eNOS) coupled to heat shock protein 90 (HSP90) with tetrahydrobiopterin (BH₄) as a cofactor with adequate supply of arginine as substrate. The eNOS protein is bound to caveolin-1 (Cav-1) prior to its release by a calcium-calmodulin (CaM) dependent process. Endothelin acting through endothelin-B receptor (ET_B) on the endothelium stimulates NO production. Manganese superoxide dismutase (MnSOD or SOD-2) is present in the mitochondria and scavenges superoxide anions. Extracellular superoxide dismutase (ecSOD) limits the interaction (and inactivation) of NO with superoxide anions in the endothelial-smooth muscle interface. NO reaches the smooth muscle cell and binds to reduced soluble guanylate cyclase (sGC) which in turn catalyzes the conversion of GTP to cGMP. Cyclic GMP is an important second messenger that reduces the cytosolic concentration of ionic calcium [Ca⁺⁺]_i. Reduced concentration of ionic calcium leads to dephosphorylation of myosin light chains (MLC) resulting in smooth muscle relaxation. In PPHN, endothelial dysfunction leads to uncoupling of eNOS. Low levels of MnSOD and possibly ecSOD increase oxidative stress and formation of superoxide anions. Superoxide anions inactive nitric oxide resulting in the formation of toxic peroxynitrite. Oxidized sGC cannot be activated by NO to produce cGMP. Superoxide anions stimulate phosphodiesterase 5 (PDE5) activity and enhance breakdown of cGMP. Pulmonary arterial endothelial cells from PPHN pulmonary arteries produce increased levels of endothelin-1 (ET-1), a powerful pulmonary vasoconstrictor. ET-1 acts through ET_A receptor and stimulates Rho-A, Rho-kinase (ROCK) pathway leading to phosphorylation of MLC and smooth muscle contraction. The pulmonary arterial endothelial cells have low levels of ET_B receptors. The net effect is reduced cGMP (vasodilator second messenger) and sensitization of the smooth muscle to ionic calcium. (Copyright-Lakshminrusimha and Steinhorn). Modified from Polin and Fox – Fetal and Neonatal Physiology 5^th Edition

Figure 5.. Nitrosative stress and the role of inhaled nitric oxide and inspired oxygen.

3-NT staining of lung sections from lambs with PPHN induced by antenatal ligation of the ductus arteriosus and ventilated for 24 hours.

1. Lambs were ventilated with 100% oxygen for 24 hours irrespective of PaO₂ levels.

2. Lambs were ventilated with 100% oxygen and 20 ppm iNO for 24 hours irrespective of PaO₂ levels.

3. Lambs were ventilated with 100% oxygen and 20 ppm iNO for 24 hours irrespective of PaO₂ levels. These lambs received a dose of intratracheal recombinant human superoxide dismutase (rhSOD) mixed with surfactant at birth.

4. Lambs were ventilated with titrated inspired oxygen to maintain preductal PaO₂ between 50 and 80 mmHg and 20 ppm iNO for 24 hours.

Modified from reference #

Figure 6.

Role of prenatal and postnatal oxidative stress in the pathogenesis of bronchopulmonary dysplasia (BPD) and pulmonary hypertension (PH). Growth restriction, both prenatal (IUGR) and postnatal (PNGR) contribute to development of PH. Cord blood placental growth factor (PIGF), granulocyte-colony stimulating factor (G-CSF), and vascular endothelial growth factor-A (VEGF-A) Copyright Satyan Lakshminrusimha

Figure 7.

Recent randomized controlled trials in extremely preterm infants with interventions that could potentially reduce oxidative stress and their outcomes. Some of these trials were stopped early due to negative outcomes in the intervention group. The Australian placental transfusion trial [88], Oei et al [90], Katheria et al [89], NeOProM trials [91] and the SAIL randomized control trial [92] are highlighted in this figure. The patient population studied, intervention, primary outcome and negative findings in subgroups (if any) are shown in boxes. Copyright Satyan Lakshminrusimha