Whole blood hypoxia-related gene expression reveals novel pathways to obstructive sleep apnea in humans

doi:10.1016/j.resp.2013.08.012

Respiratory Physiology & Neurobiology

Volume 189, Issue 3, 1 December 2013, Pages 649-654

https://doi.org/10.1016/j.resp.2013.08.012 Get rights and content

Highlights

•
Processes of preconditioning in response to hypoxia could initiate during mild-moderate apnea.
•
A total of 13 genes were identified in severe OSA patients.
•
PRPF40A and PLOD3 gene expressions were strongly and independently associated with AHI scores.

Abstract

In this study, our goal was to identify the key genes that are associated with obstructive sleep apnea (OSA). Thirty-five volunteers underwent full in-lab polysomnography and, according to the sleep apnea hypopnea index (AHI), were classified into control, mild-to-moderate OSA and severe OSA groups. Severe OSA patients were assigned to participate in a continuous positive airway pressure (CPAP) protocol for 6 months. Blood was collected and the expression of 84 genes analyzed using the RT² Profiler™ PCR array. Mild-to-moderate OSA patients demonstrated down-regulation of 2 genes associated with induction of apoptosis, while a total of 13 genes were identified in severe OSA patients. After controlling for body mass index, PRPF40A and PLOD3 gene expressions were strongly and independently associated with AHI scores. This research protocol highlights a number of molecular targets that might help the development of novel therapeutic strategies.

Introduction

Obstructive sleep apnea (OSA) is characterized by repetitive obstruction of the upper airway, resulting in pauses in breathing and subsequent oxygen desaturation. The majority of sleep disorders, including OSA, are caused by complex interactions between genes and the environment, leading studies to focus on the extent to which genes predetermine susceptibility to intermittent apneas, as well as on the effects of hypoxia and sleep fragmentation on the expression of candidate genes (Arnardottir et al., 2009). Previous studies have identified a key area of genetic influence as the cellular reaction to hypoxia (Arnardottir et al., 2009). Knowledge of the genetic factors underlying the phenotypic traits following hypoxia insults can reveal how cells confer vulnerability or resistance to hypoxia and modulate responsiveness to therapeutic interventions aimed at restoring respiratory, cardiovascular, cognitive, and metabolic functions (Cassavaugh and Lounsbury, 2011). Here, our goal was to identify the genes that are associated with hypoxia that play a role in OSA. This approach resulted in the identification of novel candidate genes of OSA, pointing the way to how genetic factors may modulate apneic susceptibility, as well as help the potential development of novel therapeutic targets.

Section snippets

Study population and polysomnography

A total of 35 volunteers underwent full in-lab polysomnography (PSG; EMBLA System N7000, software RemLogic Version 2.0, CO, USA) during their habitual sleep time. Standard montage and criteria were used for scoring sleep stages, arousals, leg movements, and respiratory events according to the guidelines from the American Academy of Sleep Medicine, including the recommend rule for hypopneas – a decrease of ≥30% in the nasal pressure signal of baseline, with a duration ≥10 s, with a desaturation

Sleep pattern – evaluation of apnea-hypopnea index

The control, mild-to-moderate, and severe OSA groups were comparable in age (Control 46.4 ± 2.6; OSA 5 to <30 41.7 ± 2.9; OSA >30 46.2 ± 2.3 years), sleep efficiency, percentage spent in NREM sleep stage N1, and REM sleep; however, the BMI was higher in the severe OSA group (Control 26.1 ± 0.7; OSA 5 to <30 26.6 ± 0.7; OSA >30 32.2 ± 1.4 Kg/m²; p < 0.05). The AHI was 2.4 events/hour in the control group, 13.2 in the patients with mild-to-moderate OSA, and 56.6 events/hour in the patients with severe OSA (p <

Discussion

Here we show direct correlation of physiological and molecular markers for OSA. The comparison between mild-to-moderate and severe OSA patients to healthy individuals showed down-regulation of DAPK3 and KAT5 genes, two molecules associated with induction of apoptosis. Taken together, we propose that the regulation of these key genes by OSA may, in part, underlie preconditioning. Future studies should examine this possibility. Moreover, our data identified independent and strong associations

Conclusions

Here, we propose for the first time integration between the molecular and physiological responses following a hypoxia insult within a network inference framework: a gene toolbox. In developing such a framework, we endeavor to define checkpoints spanning clinical application, in an attempt to help the design of novel strategies in the search for effective targets to treat OSA.

Authors’ contribution

(1) JCP, LB, SG, and DRM performed the conception and design of the study, or acquisition of data, or analysis and interpretation of data; (2) JCP, CG, LB, SG, DRM and ST involved in drafting the article or revising it critically for important intellectual content; and (3) JCP, CG, LB, SG, DRM, and ST involved final approval of the version to be submitted.

Conflict of interest

The authors declare that there is no conflict of interest.

Acknowledgements

This work was supported by grants from Associação Fundo de Incentivo à Pesquisa (AFIP), CNPq (#558924/2008-5 to JCP, CG, LRAB, ST) and FAPESP (#98/14303-3 to ST). We are grateful to Leiko K. Zanin for contacting patients and for verifying the adherence of CPAP treatment, Dr. Altay Alves Lino de Souza for statistical assistance, and all the efforts of Renata Pellegrino and Diva Maria Lima in helping the collection of data.

References (10)

M.J. Carrozza et al.
The diverse functions of histone acetyltransferase complexes
Trends Genet.
(2003)
J. Heikkinen et al.
Dimerization of human lysyl hydroxylase 3 (LH3) is mediated by the amino acids 541–547
Matrix Biol.
(2011)
L. Lavie et al.
Ischemic preconditioning as a possible explanation for the age decline relative mortality in sleep apnea
Med. Hypotheses
(2006)
J.M. Marin et al.
Long-term cardiovascular outcomes in men with obstructive sleep apnoea-hypopnoea with or without treatment with continuous positive airway pressure: an observational study
Lancet
(2005)
S. Tufik et al.
Obstructive sleep apnea syndrome in the Sao Paulo Epidemiologic Sleep Study
Sleep Med.
(2010)

There are more references available in the full text version of this article.

Cited by (13)

Chronic intermittent hypoxia impaired collagen synthesis in mouse genioglossus via ROS accumulation: A transcriptomic analysis
2023, Respiratory Physiology and Neurobiology
Citation Excerpt :
We found several significant DEGs, such as Col1a1, Col1a2, Mmp2, Col3a1, Col5a1, Fmod, Col5a2, Tgfb3, Ctsk, Col12a1, and P4ha1. Perry et al. (2013) found independent and strong associations between the apnea-hypopnea index and PLOD3 gene expression; PLOD3 is a multifunctional enzyme of collagen biosynthesis that catalyzes the posttranslational modifications of lysine in collagen and collagen-like proteins (Perry et al., 2013). Su et al. (2020) reported that bladder detrusor muscle bundles were disordered and loose and that the content of collagen fibers was reduced in CIH rats (Su et al., 2020).
Obstructive sleep apnea (OSA) is a sleep-related breathing disorder characterized by intermittent and recurrent upper airway collapse during sleep that leads to chronic intermittent hypoxia (CIH). The genioglossus (GG) is the largest dilator muscle, which controls the upper airway and plays an important role in OSA pathology. Elucidating its genetic alterations may help identify potential targets for OSA. However, the genetic aspects of the GG in CIH mice remain unclear. Here, we have conducted an RNA sequencing (RNA-Seq) analysis to assess the differentially expressed genes (DEGs) in the GG between CIH mice and normoxia (NOR) mice. A total of 637 DEGs were identified to be dysregulated in CIH mice compared with control mice. Bioinformatics analysis showed that the DEGs were related to various physiological processes, such as the endogenous stimulus responses, cellular component organization and metabolic processes. Extracellular matrix (ECM)–receptor interaction was the top KEGG pathway in the environmental information processing category with high significance and large fold changes. From the gene weight distributions of collagen (Col)-related biological processes (BPs), we found several significant DEGs, such as Col1a1, Col1a2, Mmp2, Col3a1, Col5a1, Fmod, and Col5a2. A PPI network showed that Col1a1 was linked to ECM–receptor interactions, responses to reactive oxygen species (ROS) and Col-related BPs. It was verified in vivo and in vitro that hypoxia can induce excess ROS and reduce Col expression levels. Moreover, we found NAC can effectively scavenge ROS and restore collagen synthesis. These findings contribute to a better understanding of the mechanisms linking OSA and upper airway muscle injury and may help identify potential therapeutic targets.
Whole blood gene expression signature in patients with obstructive sleep apnea and effect of continuous positive airway pressure treatment
2021, Respiratory Physiology and Neurobiology
Citation Excerpt :
On a gene expression level, OSA appears to be a complex disease with multiple genes and pathways being affected and the common association of OSA and obesity further complicates the search for true OSA biomarkers (Arnardottir et al., 2009). During the last decade, there have been series of reports published based on analysis of either whole blood or peripheral blood mononuclear cells transcriptome using a microarray approach comparing OSA patients to controls (Chen et al., 2017; Hoffmann et al., 2007; Lin et al., 2014), the effect of continuous positive airway pressure (CPAP) treatment (Gharib et al., 2014; Peng et al., 2019; Perry et al., 2013) or healthy volunteers exposed to the intermittent hypoxia (Polotsky et al., 2015). These studies have generated dissimilar results with up- or down-regulation of different genes involved in a variety of biological pathways, such as systemic and vascular inflammation, neoplastic processes, induction of apoptosis, and cell adhesion and communication
The molecular mechanisms of obstructive sleep apnea (OSA), in particular the gene expression patterns in whole blood of patients with OSA, can shed more light on the underlying pathophysiology of OSA and suggest potential biomarkers. In the current study, we have enrolled thirty patients with untreated moderate-severe OSA together with 20 BMI, age, and sex-matched controls and 15 normal-weight controls. RNA-sequencing of whole blood and home sleep apnea testing were performed in the untreated state and after three and twelve months of continuous positive airway pressure (CPAP) treatment. Analysis of the whole blood transcriptome of the patients with OSA revealed a unique pattern of differential expression with a significant number of downregulated immune-related genes including many heavy and light chain immunoglobulins and interferon-inducible genes. This was confirmed by the gene ontology analysis demonstrating enrichment with the biological processes associated with various immune functions. Expression of these genes was recovered after three months of CPAP treatment. After 12 months of CPAP treatment, the overall gene expression profile returns to the initial, untreated level. In addition, we have confirmed the importance of choosing BMI-matched controls as a reference group as opposed to normal-weight healthy individuals based on the significantly different gene expression signatures between these two groups.
Low oxygen saturation during sleep reduces CD1D and RAB20 expressions that are reversed by CPAP therapy
2020, EBioMedicine
Citation Excerpt :
Intermittent hypoxemia has been shown to affect gene expression in peripheral blood mononuclear cells of healthy volunteers [10]. Six studies ranging in size from 18 to 48 individuals have evaluated the impact of OSA on gene expression in peripheral blood monocytes or total leukocytes, either cross-sectionally in comparison to those without OSA [10–12], or pre- and post-treatment with continuous positive airway pressure (CPAP; [13,14]), the most common therapy for OSA, or following two weeks of CPAP withdrawal into either sham or oxygen supplement treatment [15]. In addition to their small sample sizes, some of these studies were limited by targeting a specific patient population (e.g., children; [12]), a limited set of genes [11], and by a focus on dichotomized (presence/absence) OSA traits.
Sleep Disordered Breathing (SDB) is associated with a wide range of pathophysiological changes due, in part, to hypoxemia during sleep. We sought to identify gene transcription associations with measures of SDB and hypoxemia during sleep, and study their response to treatment.
In two discovery cohorts, Framingham Offspring Study (FOS; N = 571) and the Multi-Ethnic Study of Atherosclerosis (MESA; N = 580), we studied gene expression in peripheral blood mononuclear cells in association with three measures of SDB: Apnea Hypopnea Index (AHI); average oxyhemoglobin saturation (avgO2) during sleep; and minimum oxyhemoglobin saturation (minO2) during sleep. Associated genes were used for analysis of gene expression in the blood of 15 participants with moderate or severe obstructive sleep apnea (OSA) from the Heart Biomarkers In Apnea Treatment (HeartBEAT) trial. These genes were studied pre- and post-treatment (three months) with continuous positive airway pressure (CPAP). We also performed Gene Set Enrichment Analysis (GSEA) on all traits and cohort analyses.
Twenty-two genes were associated with SDB traits in both MESA and FOS. Of these, lower expression of CD1D and RAB20 was associated with lower avgO2 in MESA and FOS. CPAP treatment increased the expression of these genes in HeartBEAT participants. Immunity and inflammation pathways were up-regulated in subjects with lower avgO2; i.e., in those with a more severe SDB phenotype (MESA), whereas immuno-inflammatory processes were down-regulated following CPAP treatment (HeartBEAT).
Low oxygen saturation during sleep is associated with alterations in gene expression and transcriptional programs that are partially reversed by CPAP treatment.
An integrated multi-omics analysis of sleep-disordered breathing traits implicates P2XR4 purinergic signaling
2023, Communications Biology
Epigenetics of obstructive sleep apnea syndrome: a systematic review
2021, Journal of Clinical Sleep Medicine
Lumbar spine abnormalities in patients with obstructive sleep apnoea
2021, Scientific Reports

View all citing articles on Scopus

View full text

Short communicationWhole blood hypoxia-related gene expression reveals novel pathways to obstructive sleep apnea in humans

Highlights

Abstract

Introduction

Section snippets

Study population and polysomnography

Sleep pattern – evaluation of apnea-hypopnea index

Discussion

Conclusions

Authors’ contribution

Conflict of interest

Acknowledgements

Trends Genet.

Matrix Biol.

Med. Hypotheses

Lancet

Sleep Med.

Short communication
Whole blood hypoxia-related gene expression reveals novel pathways to obstructive sleep apnea in humans