EGLN1 variants influence expression and SaO2 levels to associate with high-altitude pulmonary oedema and adaptation

Abstract EGLN1 [encoding HIF (hypoxia-inducible factor)-prolyl hydroxylase 2] plays a pivotal role in the HIF pathway and has emerged as one of the most intriguing genes with respect to physiology at HA (high altitude). EGLN1, being an actual oxygen sensor, appears to have a potential role in the functional adaptation to the hypobaric hypoxic environment. In the present study, we screened 30 polymorphisms of EGLN1, evaluated its gene expression and performed association analyses. In addition, the role of allelic variants in altering TF (transcription factor)-binding sites and consequently the replacement of TFs at these loci was also investigated. The study was performed in 250 HAPE-p [HAPE (HA pulmonary oedema)-patients], 210 HAPE-f (HAPE-free controls) and 430 HLs (healthy Ladakhi highland natives). The genotypes of seven polymorphisms, rs1538664, rs479200, rs2486729, rs2790879, rs480902, rs2486736 and rs973252, differed significantly between HAPE-p and HAPE-f (P < 0.008). The genotypes AA, TT, AA, GG, CC, AA and GG of rs1538664, rs479200, rs2486729, rs2790879, rs480902, rs2486736 and rs973252, prevalent in HAPE-p, were identified as risk genotypes and their counterpart homozygotes, prevalent in HLs, were identified as protective. EGLN1 expression was up-regulated 4.56-fold in HAPE-p (P = 0.0084). The risk genotypes, their haplotypes and interacting genotypes were associated with up-regulated EGLN1 expression (P < 0.05). Similarly, regression analysis showed that the risk alleles and susceptible haplotypes were associated with decreased SaO 2 (arterial oxygen saturation) levels in the three groups. The significant inverse correlation of SaO 2 levels with PASP (pulmonary artery systolic pressure) and EGLN1 expression and the association of these polymorphisms with SaO 2 levels and EGLN1 expression contributed to uncovering the molecular mechanism underlying hypobaric hypoxic adaptation and maladaptation

Additional file 1 of Differential methylation in EGLN1 associates with blood oxygen saturation and plasma protein levels in high-altitude pulmonary edema

Additional file 1. Fig. S1 Levels of SpO2 % in HAPE-p, HAPE-f and HLs. Fig. S2 Dot plot of CpG methylation in EGLN1 in the three study groups, i.e., HAPE-p, HLs and HAPE-f. It revealed 97 CpG sites in EGLN1 CpG island 179. Fig. S3 Dot plot of CpG methylation in HIF1AN in the three study groups, i.e., HAPE-f, HAPE-p and HLs. It revealed 46 CpG sites in HIF1AN CpG island 47. Table S1 Methylation distribution of CpG sites of EGLN1 and HIF1AN in each subject of the three study groups i.e., HAPE-f, HAPE-p and HLs. Table S2 Real-time PCR conditions for EGLN1 and HIF1AN. Table S3 Sodium bisulfite-conversion-based methylation PCR Primers and conditions for EGLN1 and HIF1AN

The EDN1 Missense Variant rs5370G > T Regulates Adaptation and Maladaptation under Hypobaric Hypoxia

Endothelin 1 (EDN1) encodes a potent endogenous vasoconstrictor, ET1, to maintain vascular homeostasis and redistribution of tissue blood flow during exercise. One of the EDN1 missense polymorphisms, rs5370 G/T, has strongly been associated with cardiopulmonary diseases. This study investigated the impact of rs5370 polymorphism in high-altitude pulmonary oedema (HAPE) disorder or maladaptation and adaptation physiology in a well-characterized case–control study of high-altitude and low-altitude populations comprising 310 samples each of HAPE-patients, HAPE-free controls and native highlanders. The rs5370 polymorphism was genotyped, and the gene expression and plasma level of EDN1 were evaluated. The functional relevance of each allele was investigated in the human embryonic kidney 293 cell line after exposure to hypoxia and computationally. The T allele was significantly more prevalent in HAPE-p compared to HAPE-f and HLs. The EDN1 gene expression and ET1 bio-level were significantly elevated in HAPE-p compared to controls. Compared to the G allele, the T allele was significantly associated with elevated levels of ET-1 in all three study groups and cells exposed to hypoxia. The in silico studies further confirmed the stabilizing effect of the T allele on the structural integrity and function of ET1 protein. The ET1 rs5370 T allele is associated with an increased concentration of ET-1 in vivo and in vitro, establishing it as a potent marker in the adaptation/maladaptation physiology under the high-altitude environment. This could also be pertinent in endurance exercises at high altitudes

Interactions among Vascular-Tone Modulators Contribute to High Altitude Pulmonary Edema and Augmented Vasoreactivity in Highlanders

<div><h3>Background</h3><p>The interactions among various biomarkers remained unexplored under the stressful environment of high-altitude. Present study evaluated interactions among biomarkers to study susceptibility for high altitude pulmonary edema (HAPE) in HAPE-patients (HAPE-p) and adaptation in highland natives (HLs); both in comparison to HAPE-free sojourners (HAPE-f).</p> <h3>Methodology/Principal Findings</h3><p>All the subjects were recruited at 3500 m. We measured clinical parameters, biochemical levels in plasma and gene expression using RNA from blood; analyzed various correlations between and among the clinical parameters, especially arterial oxygen saturation (SaO₂) and mean arterial pressure (MAP) and biochemical parameters like, asymmetric dimethylarginine (ADMA), serotonin (5-HT), 8-iso-prostaglandin F2α (8-isoPGF2α), endothelin-1 (ET-1), plasma renin activity (PRA), plasma aldosterone concentration (PAC), superoxide dismutase (SOD) and nitric oxide (NO) in HAPE-p, HAPE-f and HLs. ADMA, 5-HT, 8-isoPGF2α, ET-1 levels, and PAC were significantly higher (p<0.0001, each), whereas SOD activity and NO level were significantly lower in HAPE-p than HAPE-f (p≤0.001). Furthermore, ADMA, 5-HT, 8-isoPGF2α, NO levels and PAC were significantly higher (p<0.0001), whereas ET-1 level significantly (p<0.0001) and SOD activity non-significantly (p>0.05) lower in HLs than HAPE-f. The expression of respective genes differed in the three groups. In the correlations, SaO₂ inversely correlated with ADMA, 5-HT and 8-isoPGF2α and positively with SOD in HAPE-p (p≤0.009). MAP correlated positively with 5-HT and 8-isoPGF2α in HAPE-p and HLs (p≤0.004). A strong positive correlation was observed between ADMA and 5-HT, 5-HT and 8-isoPGF2α (p≤0.001), whereas inverse correlation of SOD with ET-1 in HAPE-p and HLs (p≤0.004), with 5-HT and 8-isoPGF2α in HAPE-p (p = 0.01) and with 5-HT in HLs (p = 0.05).</p> <h3>Conclusions/Significance</h3><p>The interactions among these markers confer enhanced vascular activity in HLs and HAPE in sojourners.</p> </div

Sexual Dimorphism of Dexamethasone as a Prophylactic Treatment in Pathologies Associated With Acute Hypobaric Hypoxia Exposure

Dexamethasone can be taken prophylactically to prevent hypobaric hypoxia-associated disorders of high-altitude. While dexamethasone-mediated protection against high-altitude disorders has been clinically evaluated, detailed sex-based mechanistic insights have not been explored. As part of our India-Leh-Dexamethasone-expedition-2020 (INDEX 2020) programme, we examined the phenotype of control (n = 14) and dexamethasone (n = 13) groups, which were airlifted from Delhi (∼225&nbsp;m elevation) to Leh, Ladakh (∼3,500&nbsp;m), India, for 3&nbsp;days. Dexamethasone 4&nbsp;mg twice daily significantly attenuated the rise in blood pressure, heart rate, pulmonary pressure, and drop in SaO2 resulting from high-altitude exposure compared to control-treated subjects. Of note, the effect of dexamethasone was substantially greater in women than in men, in whom the drug had relatively little effect. Thus, for the first time, this study shows a sex-biased regulation by dexamethasone of physiologic parameters resulting from the hypoxic environment of high-altitude, which impacts the development of high-altitude pulmonary hypertension and acute mountain sickness. Future studies of cellular contributions toward sex-specific regulation may provide further insights and preventive measures in managing sex-specific, high-altitude-related disorders

Clinical correlations in the three groups i.e. HAPE-p, HAPE-f and HLs.

<p>An inverse correlation was obtained between the clinical parameters viz MAP and SaO₂ in the three groups; MAP, mmHg; SaO₂, %.</p

Dexamethasone prophylaxis protects from acute high-altitude illness by modifying the peripheral blood mononuclear cell inflammatory transcriptome

Acute high-altitude (HA) exposure can induce several pathologies. Dexamethasone (DEX) can be taken prophylactically to prevent HA disease, but the mechanism by which it acts in this setting is unclear. We studied the transcriptome of peripheral blood mononuclear cells (PBMCs) from 16 subjects at low altitude (LA, 225 m) and then 3 days after acute travel to HA (3500 m) during the India-Leh-Dexamethasone-Expedition-2020 (INDEX2020). Half of the participants received oral DEX prophylaxis 4 mg twice daily in an unblinded manner, starting 1 day prior to travel to HA, and 12 h prior to the first PBMC collection. PBMC transcriptome data were obtained from 16 subjects, half of whom received DEX. The principal component analysis demonstrated a clear separation of the groups by altitude and treatment. HA exposure resulted in a large number of gene expression changes, particularly in pathways of inflammation or the regulation of cell division, translation, or transcription. DEX prophylaxis resulted in changes in fewer genes, particularly in immune pathways. The gene sets modulated by HA and DEX were distinct. Deconvolution analysis to assess PBMC subpopulations suggested changes in B-cell, T-cell, dendritic cell, and myeloid cell numbers with HA and DEX exposures. Acute HA travel and DEX prophylaxis induce significant changes in the PBMC transcriptome. The observed benefit of DEX prophylaxis against HA disease may be mediated by suppression of inflammatory pathways and changing leukocyte population distributions

Clinical characteristics of the HAPE-p, HAPE-f and HLs.

<p>Data are presented as mean ± standard deviation and are compared by Student's <i>t</i>-test. n, number of subjects; SBP, systolic blood pressure; DBP, diastolic blood pressure; MAP, mean arterial pressure; RR, respiratory rate; PR, pulse rate; SaO₂, arterial oxygen saturation; PASP, pulmonary artery systolic pressure.</p

Schematic presentation of the studied biomarkers in a physiological function under hypobaric hypoxia.

<p>Interactions of several of these biomarkers that translate into the incidence of HAPE are also interpretable through the renin–angiotensin–aldosterone system, Kinin–kallikrein system, and the pathways of ET-1, 5-HT, NO signaling and oxidative-stress. ADMA, asymmetric dimethylarginine; ATII, angiotensin II; AT-1R, angiotensin-II type I receptor; AT-2R, angiotensin-II type II receptor; AngI, angiotensin I; AngII, angiotensin II; ACE, angiotensin-I converting enzyme; AGT, angiotensinogen; 8-isoPGF2α, 8-iso-prostaglandinF2α; ROS, reactive oxygen species; SOD, superoxide dismutase; O2˙-, superoxide anion; ONOO⁻, peroxynitrite; 5-HT, serotonin; ET-1, endothelin-1; ET-_A and ET-_B, endothelin receptors A and B; NO, nitric oxide; NOS3, endothelial nitric oxide synthase; VSMC, vascular smooth muscle cell; H₂O₂, hydrogen peroxide; H₂O, water; EFA, essential fatty acids.</p