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Research on Cytochrome P450 enzymes may be the future of prevention and treatment of Oxygen and Nitric Oxide injuries in preterm and term infants
By Xanthi Couroucli, M.D.
Many scientists played key roles in developing the basic concepts in neonatal-perinatal medicine that helped revolutionize the evolving clinical care and inspired further research. Although the birth of neonatal medicine has its roots in the 19th century, only in 1960 was the term “neonatology” designated to the art and science of diagnosis and treatment of disorders of the newborn infant. Since then, we have witnessed an explosion of new information about the pathophysiologic processes involved in diseases of the neonate. Many of these disorders can be complications of a premature birth or in term infants because of pulmonary hypertension of the newborn.
Preterm birth (below 37 weeks of gestation) happens in about 10 percent of deliveries in the United States. The more premature the infant is the higher the possibility to develop hypoxic respiratory failure due to immature lungs and surfactant insufficiency (a condition that is called hyaline membrane disease or
HMD). Surfactant is a substance that is produced by the type II epithelial cells of the small airways and keeps the alveoli open. The premature lung is Surfactant deficient, and as a result the alveoli collapse, form hyaline membranes, oxygen exchange is disrupted and hypoxia ensues. The recent treatment of this life-threatening condition is supplemental oxygen, ventilation and administration of exogenous surfactant into the airways of the preterm infant. Despite the surfactant administration and many advances in the field of perinatal-neonatal medicine, the sickest and most immature infants who are surviving more frequently now develop more severe and prolonged complications of prematurity.
My research has been focusing on two of these complications of prematurity, the chronic lung disease (CLD) of the preterm infants, or bronchopulmonary dysplasia (BPD) and more recently, retinopathy of prematurity (ROP) as well as the pathophysiology of the
persistent pulmonary hypertension of the newborn (PPHN).
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This
represents lung sections of 2 days old rats, in room air
(A), heperoxia (B), and hyperoxia and Nitric Oxide-40ppm(C).
The
arrows show that when antibody to CYP1A1 was used (immunohistochemistry),
the epithelial cells of the airways and endothelial cells of
the pulmonary vessels expressed this gene.
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Bronchopulmonary dysplasia is defined as the continued need for supplemental oxygen beyond 28 days of life or 36 weeks of gestational age. The spectrum of BPD ranges from infants with minimal radiographic changes associated with slight cardiorespiratory distress to disabling cardiorespiratory failure seen after prolonged ventilation with high ventilatory pressures and inspired oxygen. Despite all the scientific advancements we still find that about 20-50 percent of the preterm infants below 32 weeks of gestation may develop BPD. BPD is the clinical evolution of a sequence of injuries involving a spectrum of factors. Among these factors, the interface of mechanical ventilation and oxygen with the lung of the vulnerable newborn plays
a major role. Infections (bacterial or viral) also had been established as a major predisposing factor to BPD. Indeed, in 1999 in a collaborative study with Dr. Jeff Towbin’s laboratory, we found that preterm infants from Texas Children’s Hospital had higher possibility to develop BPD if they had congenital adenoviral infection. Oxygen administration is an essential component of the care of these neonates. More recently, the addition of inhaled Nitric Oxide for its vasodilatory and anti-inflammatory effects is considered for the patients with severe BPD. Clinical and laboratory studies about the effects of combined Nitric Oxide and hyperoxia on the developing immature lung are still on the way. On the other hand, according to many studies, pulmonary tissue appears particularly sensitive to the toxic effects of oxygen to which its cells are directly exposed. It is well established by now, that the two major manifestations of oxygen toxicity involve damage to the lungs and immature retina.
More than 80 percent of premature babies weighing less than 1,000 grams develop some degree of
ROP, the incidence of which is rising as a result of the improved survival of such infants. After birth, exposure to higher oxygen concentrations causes the retina to “read” that oxygen delivery is excessive. As a result, the developing retinal capillaries constrict and disintegrate, decreasing oxygen delivery. As the retina matures, it becomes increasingly metabolically active and, as a result of an inadequate blood supply, progressively more hypoxic. Retinal abnormal neovascularization is then stimulated in response to the hypoxic environment. This results in bleeding and retinal detachment, which is the major cause of blindness in ROP.
Oxygen administration is also absolutely essential treatment for term or near-term infants with hypoxic respiratory failure due to
persistent pulmonary hypertension of the newborn (PPHN). The incidence could reach two out of 1,000 live term births. These infants can be critically ill with high oxygen requirements and ventilatory settings and, in many cases, the addition of other therapeutic modalities like inhaled Nitric Oxide or even extracorporeal membrane oxygenation
(ECMO) is necessary. PPHN is a disorder of the pulmonary vascular bed that is either maladapted or maldeveloped. Transition from intrauterine to extrauterine life requires the pulmonary vascular resistance of the fetus to decrease abruptly. In infants with PPHN, this decrease does not happen. Although these infants do not develop ROP, some of them do require prolonged administration of oxygen (more than 28 days) and develop chronic lung disease. Therefore, oxygen toxicity could still affect the developing lungs of even the term newborns. Another phenomenon that has not been studied a lot is the possible toxicity, at the molecular level, of the combined administration of inspired oxygen and inhaled nitric oxide in these patients. Nitric oxide is a short-acting inhaled gas that acts on the endothelium of the pulmonary vessels and causes vasodilation. Studies have shown that the addition of low doses of inhaled nitric oxide to the high oxygen treatment in babies with PPHN had reduced the need for ECMO.
As we can see from the above summary, the use of supplemental oxygen in our newborns is routinely needed for the treatment of hypoxic respiratory failure, which is frequently encountered in preterm and term infants. However, despite the fact that oxygen administration may be life saving, hyperoxic therapy may contribute to tissue damage and the development of BPD and ROP. For years now, researchers have been trying to reduce and prevent these toxicities by studying the role of various antioxidant systems in animal and cell models under hyperoxic conditions.
Exposure of experimental animals to hyperoxia causes lung and retinal injury. The data are conflicting on the issue of whether administration of inhaled nitric oxide is protective or damaging in the presence of hyperoxia in lung cells and animal models.
Cytochrome P450 (CYP) enzymes may be the answer
CYP enzymes belong to a superfamily of hemoproteins that play important roles in the metabolism of exogenous and endogenous chemicals, steroids and hormones. In humans there are 57 different gene families in tissues of the body. These enzymes, including the more prominent CYP1A1 and 1A2, have also been implicated in the formation and further reactions of reactive oxygen species (ROS) and may play a role in pulmonary and other tissues’ oxygen toxicity. The CYP enzymes are developmentally regulated. Seventy percent of the CYP are expressed at one stage or another during development of animals and humans. Also, these enzymes are inducible during prenatal and postnatal age by various substances including carcinogens, antibiotics and derivatives of vitamins like b-naphthoflavone. Nitric oxide appears to modulate the expression of CYP enzymes, as well. Studies have shown that nitric oxide can downregulate the expression of CYP enzymes, but the biological significance of this has not yet been demonstrated.
Taken all together, what we know from our clinical experience and scientific experiments has led us to the conclusion that although the administration of supplemental oxygen and inhaled nitric oxide are life-saving treatments, they do – and can – have severe and debilitating toxicities.
The reduction of these toxicities has always been my aim in life. Inspired by my
suffering patients and my excellent mentors (Dr. Bhagavatula Moorthy from Baylor
College of Medicine and Dr. Ferid Murad from the University of Texas Houston Medical School), I started research at the molecular level that would identify pathways through which we could prevent and/or treat these toxicities. Maybe one of the major roles of these Cytochrome P450 enzymes that had been present in our cells for millions of years is to detoxify various compounds, which could play a protective role against these toxicities. If that is so, then maybe there is a way or a substance we can give to our patients to increase the expression of these enzymes and help reduce hyperoxic and nitric oxide toxic side effects.
For the last few years, we have been conducting experiments in our laboratory to test the hypothesis that CYP1A1 and 1A2 play a role in hyperoxic and nitric oxide injuries in both adult and newborn rodents and knock-out mice (mice that lack a gene). To date, we have found:
1. CYP1A1 (which is found also in the lung) most probably plays a protective role in hyperoxic lung injury of adult rats.
2. CYP1A2 (which is found in the liver but not the lungs) plays an important role in hyperoxic lung injury of adult mice.
We have found that adult mice that lack the 1A2 gene, after exposure to
hyperoxia, die of acute respiratory distress.
3. AHR (the receptor that modulates the expression of the 1A1 and 1A2 enzymes) plays a role in hyperoxic lung injury of adult mice that lack the AHR gene.
4. CYP1A2 plays an important role in hyperoxic lung and retinal injuries in newborn mice.
We have found that newborn mice that lack the 1A2 gene, after exposure to
hyperoxia, have severe retinopathy and die of acute respiratory failure.
5. CYP1A1 most probably plays an important protective role in the oxygen-induced abnormal maturation of the lungs of the newborn rats and can modulate the effects of retinoic acid and its effects on development.
6. CYP1A1 most probably plays a protective role for lung toxicity and maturation in prolonged exposures of combined hyperoxia and low doses of inhaled nitric oxide in newborn rats.
7. Finally, and very excitingly, we found that administration of b-naphthoflanes (a vitamin B derivative), which induced CYP1A1 expression in the lungs of adult rats, protected them from hyperoxic lung injury.
So, how can we take all these exciting findings from the laboratory back to our patients and help them more? Well, until now we have found that at least one family of enzymes, the CYP1A, plays most probably a protective role against these toxicities. We also have found that a vitamin-B derivative can increase the expression of these enzymes and decrease the lung hyperoxic toxicities in adult animals. Could it be that we will find the same results in our newborn animals and
in finally our patients? We all look at the future with great hope that this research field will contribute significantly to the reduction of these frequent and severe complications of necessary treatments to our newborn patients. The financial, social and physical consequences of newborn diseases to our society are so major,
that research
for their prevention and alleviation has to carry on. After all, the children are the future of every country and healthier children can generate a better future.
Xanthi Couroucli, M.D., is a member of the Texas Children’s medical staff and an assistant professor in the Department of Pediatrics at Baylor College of Medicine.
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