Molecular Basis of Hypoxia Induced Excessive Erythrocytosis

Abstract

ObjectiveSince polycythemia is a predominant trait in some high altitude dwellers (Chronic Mountain Sickness, CMS, or Monge's disease) but not others living at the same altitude in the Andes, we took advantage of this human “experiment in nature” and studied both populations (with CMS and without, non‐CMS). Although polycythemia could be advantageous at high altitude because it increases O2‐carrying capacity, this adaptive pattern to high altitude has deleterious effects since blood increases its viscosity and induces serious morbidities, such as myocardial infarction and stroke in young adults.MethodsIn order to understand the molecular basis for polycythemia of high altitude, we generated a disease in‐the dish‐model by re‐programming fibroblasts from CMS and non‐CMS subjects. In the past, we have done whole genome sequencing of subjects from this region (CMS and non‐CMS) to study selective sweeps and genetic basis of CMS. In this study, by manipulating expression of some of these genes in using our in‐vitro model system, we are delineating the functional basis and the molecular mechanism(s) linked to the pathology.ResultsAs compared to sea level controls and non‐CMS subjects who responded to hypoxia by increasing their RBCs modestly or not at all, CMS cells increased theirs remarkably (up to 60 fold) with a dose‐dependent response to graded hypoxia (1.5, 5, 10% O2). We knocked down SENP1 (a desumoylase) in CMS iPS cells using lentiviral constructs and observed a striking reduction (>90%) of the CMS excessive erythropoietic response to low O2. And, by over‐expressing SENP1 in non‐CMS iPS cells, the hypoxic response in these subjects increased enormously. By further analyzing RBC differentiation in hypoxia, we found that VEGF, GATA1 and Bcl‐xL increased their gene expression in the CMS erythroid cells, in contrast to their minimal expression in the other two populations. We demonstrate also that, by utilizing a SUMO‐GATA1 fused construct, GATA1 desumoylation, a target of SENP1, is critical for the CMS phenotype. Unlike GATA1, the over‐expression of the anti‐apoptotic gene Bcl‐xL (a GATA1 target), only partially rescued the blunted erythroid response to hypoxia in non‐CMS cells.ConclusionWe conclude that the increased erythropoietic sensitivity to hypoxia in CMS subjects is genetic in nature and depends on an increase in SENP1 expression and its desumoylation mediated activation of GATA1 under hypoxia. By combining the iPS technology with this unique Andean population that has adapted (or mal‐adaptated) to chronic hypoxia over thousands of years, we have built an important in‐vitro model. We believe that utility of this model lies not only for studying hypoxia‐induced polycythemia but also other hypoxia‐driven diseases experienced at sea level.Support or Funding InformationThis study is funded by National Institutes of Health (NIH) grants (1P01HL098053 and 5P01HD32573) to G.G. Haddad