Mitochondrial DNA 7908–8816 region mutations in maternally inherited essential hypertensive subjects in China

Essential hypertension (EH) is a common chronic disease which is becoming an urgent public health issue worldwide, accounting for 9.4 million deaths each year [1]. EH is characterized by an elevation in arterial pressure and is a major risk factor for many common causes of morbidity and mortality including myocardial infarction, congestive heart failure, stroke and kidney failure in many segments of the population [2]. EH results from the interaction between environmental and inherited risk factors, which can be caused by single-gene or multifactorial conditions.

A family history of hypertension means that individuals are more likely to suffer hypertension [3, 4]. Maternally inherited essential hypertension (MIEH) is EH that shows a pattern of maternal inheritance and is occasionally observed in the clinic [5]. mtDNA can lead to mitochondrial diseases that are exclusively transmitted from the mother. mtDNA mutations have been identified in some pathogenic diseases such as myoclonic epilepsy, lactic acidosis, mitochondrial myopathy, encephalopathy, stroke-like episodes, and ragged-red fibers or maternally inherited diabetes [6]. Accordingly, mutations in mtDNA have also been reported in MIEH [7].

Mitochondria have an inefficient DNA repair and protection system in comparison to that of nuclear DNA [8]. All the homoplasmic mtDNA mutations that have been identified as being related to MIEH have caused functional disorders. The m.4435A > G mutation that is located immediately at the 3’end of the anticodon. This location corresponds to position 37 of tRNAMet affecting codon recognition, structural formation, and stabilization of functional tRNAs [9]. The m.4263A > G mutation reduces the efficiency of the tRNAIle precursor 5′-end cleavage that is catalyzed by RNase P because it is located at the processing site for the tRNAIle 5′-end precursor [10]. The result of these mutations is abnormal mitochondrial respiration that causes oxidative stress, this uncouples the oxidative pathways for ATP synthesis, and leads to cellular energetic processes failing [11].

To date, the roles of somatic mtDNA mutations in MIEH are still poorly understood. The development of blood pressure and this increases with many factors that include the mtDNA mutation/background, nuclear genes and environmental factors [12]. There is some suggestion that gene variations are associated with hypertension; but different results have been seen in different populations [13]. The mitochondrial genome accounts for ~ 5% of the heritability of blood pressure and these increases to ~ 35% for hypertensive pedigrees [14, 15]. With improved genetic analysis techniques, in particular genome-wide association studies, genes can now be identified that are likely to contribute to the development of hypertension within populations [16]. However, most genetic mutations have been identified in the nuclear genome [17]; only a few studies have focused on the investigation of the mitochondrial genome in the development of hypertension in populations. Therefore, it is obvious that understanding mtDNA sequence alteration involvement in MIEH may improve understanding of the genetic basis and pathogenesis of MIEH.

MtDNA mutations mainly distributed in the 7908–8816 region as described previously [18]. In this study, to understand more about the molecular mechanism underlying MIEH, we undertook screening of study in the mtDNA 7908–8816 region in hypertensive and normotensive subjects in a systematic and extensive manner. We explored inherited and clinical evidence to observe the relationship between the mitochondrial genome and MIEH. We decided to focus upon a Chinese Han population, as the morbidity of EH in Chinese adults is nearly 11.8% [19] but there is a limited amount of study on this racial group.

This study aimed to investigate mtDNA 7908–8816 region mutations in a Chinese population of patients with MIEH. The results showed that the patients in the MIEH group harbored more mtDNA variants than the control group. We simultaneously identified 40 different heteroplasmic or homoplasmic mutations in 4 genes: COXII, tRNA^Lys, ATP8 and ATP6. As previously reported, most pathogenic mtDNA mutations are in tRNAs [25]. Mutations in protein-encoding genes are frequently associated with ATPase dysfunction [26]. Failure in tRNA metabolism will lead to the deficiency of mitochondrial protein synthesis [27]. Defects in mitochondrial translation consequently result in a respiratory phenotype and a decrease in ATP production,may reduce the production of ROS,and subsequently have the potential role to affect the course of hypertension [28]. The mtDNA variations were mainly distributed in regions of ATP6 binding sites, and the site of highest mutation frequency was m.8414C > T. Three single base pair changes were significantly different between the MIEH and control groups, namely C8414T in ATP8, and A8701G and G8584A in ATP6. It is becoming established that mitochondrial damage and dysfunction are important factors in cardiovascular disease [29]. To our best knowledge, this is one of the first large-scale population-based systematic screens for mitochondrial mutations and their effects on MIEH in the Chinese population. Systematic study of the relationship between disease and mtDNA mutation is important to assist with our understanding of the mechanism of the mutation and its relationship to disease, but is also able to improve diagnosis, prevention and treatment of EH.

Blood glucose, blood lipids, creatinine, urea nitrogen and other biochemical abnormalities are all very closely related to the chance of developing primary hypertension [30]. In order to clarify whether mtDNA affects the biochemical indicators of the MIEH individuals, we compared and analyzed the biochemical abnormalities of the MIEH and normal individuals. Clinical examination and evaluation of all available members in this study suggested that MIEH subjects presented significantly higher values than those of non-maternal members in BMI, WC, AC, TG, LDL, FBG, UA, CR and BUN. In our study, participants with MIEH were overweight or obese compared to participants with normal blood pressure. A normal body weight (BMI 18.5–24.9 kg/m²) should be maintained for prevention and management of hypertension [31]. The occurrence and development of MIEH, can involve these factors or these factors might occur as a result of the development of MIEH, which leads to the damage or deterioration of the target organ.

Many studies, including the analysis of maternally transmitted hypertension in a large Han Chinese pedigree, have acknowledged the role of inherited mtDNA mutations in familial MIEH [32, 33]. Here, we undertook mutational analysis of the mitochondrial DNA 7908–8816 region using PCR amplification and then sequence analysis of the PCR fragments. The present experiment showed that there were more mtDNA variations in frequency and density in the MIEH patients than those who were normotensives (NT). Among these mutations, ATP6 is a hotspot for pathogenic mutations associated with MIEH. The occurrence of mtDNA mutations in these genetically unrelated subjects affected by MIEH suggests that mutations may participate in key functional development processes of EH. There are hundreds of mitochondria and thousands of mtDNAs in a mammalian cell and the close proximity of mtDNA within mitochondria with ROS generation sites means that mtDNA is vulnerable to a high level of mutation without an efficient DNA protection and repair system [18].

Mutations of mtDNA may lead to disease, and the significant determinant of their clinical outcomes are likely to correlate with the amount of mutated mtDNA [34]. Some MIEH patients in the study were part of one family branch. Among the MIEH individuals, there was a high mutation frequency and density in mtDNA m.8584G > A and m.8701A > G mutations. Specifically, an amino acid change at the m.8414C > T in the ATP8 gene (leucine to phenylalanine) shows that mtDNA variants may be able to affect the development of hypertension in China [35]. Ancestral variants of mtDNA define population-specific mtDNA lineages or haplogroups. These were first used to trace the origins of different races and allow reconstruction of the migration of humans throughout our ancient history [36]. MtDNA lineages have been shown recently to be more prone to certain disease symptoms, including type-2 diabetes, obesity and atherothrombotic cerebral infarction. Haplogroups can also protect against myocardial infarction and increase lifespan.

MtDNA mutations, including point mutations, deletions, and duplications that affect transcription and translation of mtDNA are implicated in most mitochondrial diseases [37]. Our intriguing observation is that there was a m.8273_8281del mutation in 59 MIEH patients. This 9-bp deletion polymorphism is a phylogenetic marker of studies into evolution trends and population migration [38]. Because of its location, this polymorphism could change either downstream or upstream gene expression. Some crucial structural components of the respiratory chain, such as ATP8, ATP6 are located in the downstream of 9-bp deletion polymorphism [39]. This could affect important respiratory chain structural components, including ATP8 and ATP6 that are located downstream of this polymorphism. If these genes are abnormally expressed they could change oxidative phosphorylation and influence oxidative stress levels [40].

Compared to MIEH patients without mtDNA mutations, the onset time of hypertension for patients with mtDNA mutations was significantly ahead of schedule. This is comparable to the onset time for other Chinese patients with maternally transmitted hypertension [41]. Individuals carrying mtDNA mutations develop hypertension stimulated by environmental factors more easily. Impaired mitochondrial function may contribute increased blood pressure characteristic of aging. Heteroplasmy of mtDNA is strongly associated with hypertension, with the EH mother carrying the gene mutation, and the children with the same mutation more likely to suffer from hypertension. Regarding amino-acid changes and RNAs variants the MIEH subjects harbored more variants than the controls. A possible reason for mtDNA defects pathogenesis in hypertension is that it decreases energy production, increases ROS production, resulting in oxidative stress, disrupting signal transduction, and leading to cardiovascular and renal damage, ultimately initiating hypertension [42, 43]. In consist with previous report [44], mtSNPs may affect the development of hypertension in sporadic Chinese hypertensive subjects. Some specific mtSNP within mitochondria may have potential effect in Chinese hypertensives because of their function. How mitochondrial mtSNPs and/or haplogroups interact synergistically needs to be investigated in further studies. This study was the first step in investigating the role of mitochondria in Chinese hypertensives.

This study has some limitations. One was the small sample size and the single center nature of the study. The conclusions would have been strengthened if the more people had been enrolled. We were not able to control for genetic principal components, which is typically essential in any genomic analysis. Additional limitation was that we were not including people treated with antihypertensives, etc. this would have eliminated quite a large number of people from the analysis.

In conclusion, our data convincingly demonstrated the possibility of mitochondrial mutations being involved in the pathological process of MIEH. We also identified mitochondrial genetic characteristics in MIEH individuals. Our findings may be generalizable to other China and Asian populations with a similar lifestyle. The investigation of the role of mitochondrial dysfunction in MIEH provides critical implications on the understanding and treatment of this disorder. The present research serves as a solid foundation for further study on the association between MIEH and mitochondrial dysfunction, and the cause and effect relationship in this population.