PHD2 inactivation in Type I cells drives HIF‐2α‐dependent multilineage hyperplasia and the formation of paraganglioma‐like carotid bodies

Abstract

Key points The carotid body is a peripheral arterial chemoreceptor that regulates ventilation in response to both acute and sustained hypoxia.Type I cells in this organ respond to low oxygen both acutely by depolarization and dense core vesicle secretion and, over the longer term, via cellular proliferation and enhanced ventilatory responses.Using lineage analysis, the present study shows that the Type I cell lineage itself proliferates and expands in response to sustained hypoxia.Inactivation of HIF‐2α in Type I cells impairs the ventilatory, proliferative and cell intrinsic (dense core vesicle) responses to hypoxia.Inactivation of PHD2 in Type I cells induces multilineage hyperplasia and ultrastructural changes in dense core vesicles to form paraganglioma‐like carotid bodies.These changes, similar to those observed in hypoxia, are dependent on HIF‐2α.Taken together, these findings demonstrate a key role for the PHD2–HIF‐2α couple in Type I cells with respect to the oxygen sensing functions of the carotid body. AbstractThe carotid body is a peripheral chemoreceptor that plays a central role in mammalian oxygen homeostasis. In response to sustained hypoxia, it manifests a rapid cellular proliferation and an associated increase in responsiveness to hypoxia. Understanding the cellular and molecular mechanisms underlying these processes is of interest both to specialized chemoreceptive functions of that organ and, potentially, to the general physiology and pathophysiology of cellular hypoxia. We have combined cell lineage tracing technology and conditionally inactivated alleles in recombinant mice to examine the role of components of the HIF hydroxylase pathway in specific cell types within the carotid body. We show that exposure to sustained hypoxia (10% oxygen) drives rapid expansion of the Type I, tyrosine hydroxylase expressing cell lineage, with little transdifferentiation to (or from) that lineage. Inactivation of a specific HIF isoform, HIF‐2α, in the Type I cells was associated with a greatly reduced proliferation of Type I cells and hypoxic ventilatory responses, with ultrastructural evidence of an abnormality in the action of hypoxia on dense core secretory vesicles. We also show that inactivation of the principal HIF prolyl hydroxylase PHD2 within the Type I cell lineage is sufficient to cause multilineage expansion of the carotid body, with characteristics resembling paragangliomas. These morphological changes were dependent on the integrity of HIF‐2α. These findings implicate specific components of the HIF hydroxylase pathway (PHD2 and HIF‐2α) within Type I cells of the carotid body with respect to the oxygen sensing and adaptive functions of that organ.

Marked and rapid effects of pharmacological HIF-2α antagonism on hypoxic ventilatory control

“…These responses involve a progressive increase in ventilatory sensitivity to chemoreceptive stimuli in the carotid body. Because in clinical or physiological settings, such as chronic lung disease or altitude exposure, hypoxia is generally sustained and because genetic studies have suggested an important role for HIF-2 in this setting (19,20,23,24), we first studied the effects of PT2385 on ventilatory sensitivity in animals exposed to sustained hypoxia. Male mice were treated by gavage with 3 or 10 mg/kg PT2385 (or vehicle alone) twice daily, doses that have been shown to be effective in reducing tumor size in ccRCC xenografts in mice (14).…”

“…PT2385 was commenced 24 hours before and maintained throughout the 7-day exposure of mice to hypoxia. To monitor response in which HIF, in particular HIF-2, has been implicated by genetic interventions is in the control of respiration by the carotid body (19)(20)(21). However, in common with most studies that deploy genetic inactivation technology, such studies of HIF genetic inactivation generally aim to generate complete loss of a specific HIF-α protein in a specific cell type and do not necessarily mimic dose-dependent actions of a pharmaceutical agent that is applied systemically.…”

Molecular and cellular mechanisms of the organogenesis and development of the mammalian carotid body

“…Homozygous deletion of the HIF‐1α gene causes embryonic lethality, mainly due to lack of vascularization . The HIF‐2α protein and mRNA are intensely expressed in glomus cells and endothelial cells in adult mouse carotid body . HIF‐2α immunoreactivity is also detected during maturation of the rat carotid body, that is, at E20, P0, P2, and P4 .…”

“…Endothelial cells are derivatives of the mesoderm. On the other hand, using lineage‐labeling technology of glomus cells and sustentacular cells, respectively, the others say that a large majority of increased glomus cells during hypoxia is derived from the glomus cell lineage itself and is not via transdifferentiation from sustentacular cells . Furthermore, the existence of pre‐differentiated, immature glomus cells which express TH and are capable of rapid division in response to hypoxia, has been suggested in the adult carotid body and these cells respond to sustained hypoxia …”

“…Unlike other peripheral neural organs, such as the adrenal medulla and sympathetic ganglia, the volume of the carotid body remarkably increases in response to acute and chronic hypoxia in adult animals. Therefore, neural stem cells that can differentiate into oxygen‐sensitive mature glomus cells have been suggested to exist in the carotid body . However, the origin of these stem cells remains controversial.…”