Recent Advances in Epigenetics Of Age-Related Kidney Diseases

Abstract: Renal aging has attracted increasing attention in today’s aging society, as elderly people with advanced age are more susceptible to various kidney disorders such as acute kidney injury (AKI) and chronic kidney disease (CKD). There is no clear-cut universal mechanism for identifying age-related kidney diseases, and therefore, they pose a considerable medical and public health challenge. Epigenetics refers to the study of heritable modifications in the regulation of gene expression that do not require changes in the underlying genomic DNA sequence. A variety of epigenetic modififiers such as histone deacetylases (HDAC) inhibitors and DNA methyltransferase (DNMT) inhibitors have been proposed as potential biomarkers and therapeutic targets in numerous fields including cardiovascular diseases, immune system diseases, nervous system diseases, and neoplasms. Accumulating evidence in recent years indicates that epigenetic modifications have been implicated in renal aging. However, no previous systematic review has been performed to systematically generalize the relationship between epigenetics and age-related kidney diseases. In this review, we aim to summarize the recent advances in epigenetic mechanisms of age-related kidney diseases as well as discuss the application of epigenetic modififiers as potential biomarkers and therapeutic targets in the field of age-related kidney diseases. In summary, the main types of epigenetic processes including DNA methylation, histone modifications, and non-coding RNA (ncRNA) modulation have all been implicated in the progression of age-related kidney diseases, and therapeutic targeting of these processes will yield novel therapeutic strategies for the prevention and/or treatment of age-related kidney diseases.

Keywords: epigenetics; histone modification; DNA methylation; non-coding RNA regulation; age-related kidney diseases 

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1. Introduction 

With the rapid development of the social economy and the improvement in health care, the life expectancy of the general population has continued to increase, resulting in a signifificantly increasing proportion of the elderly population [1]. According to World Health Organization (WHO) statistics, it is estimated that there will be approximately two billion people over 60 years old worldwide by 2050 [2–4]. Among all the issues caused by aging, renal aging is becoming nonnegligible. Although the aging process does not directly cause kidney disease, aging kidneys are more susceptible to various adverse factors such as high blood pressure, diabetes, obesity, or primary renal disorders, which may contribute to the development of renal pathologies. Aging is associated with an inevitable time-dependent decline in kidney function [5,6]. The main pathological characteristics of the aged kidney include arteriosclerosis, glomerulosclerosis, tubular atrophy, and interstitial fibrosis [7]. Specifically, numerous studies have indicated that aging is recognized as a major contributor to the increased incidence of acute kidney injury (AKI) and chronic kidney disease (CKD) in the overall population [7–9]. Furthermore, the incidence of deaths due to renal dysfunction is increasing globally in parallel with the aging population. However, the underlying mechanisms of renal aging have not been clearly elucidated yet, and remain the focus of current research. Thus, wisely addressing the challenges of population aging requires identifying the root causes of this phenomenon as well as effectively solving the issues, which will bring huge social benefits. 

So far, several mechanisms have been found to be closely associated with age-associated organ function decline including genome instability, telomere attrition, epigenetic alterations, loss of proteostasis, dysregulated nutrient sensing, mitochondrial dysfunction, cellular senescence, and stem cell exhaustion as well as altered intercellular communication [6,10], among which epigenetic changes are the newly identified hallmarks of aging in the past decade [6,11]. Epigenetics mainly refers to the study of molecular processes that regulate gene expression as well as phenotype without changing the primary DNA sequence, which mainly involves DNA methylation, histone post-translational modifications, chromatin remodeling, and regulation by non-coding RNAs (ncRNAs). In recent years, the critical role of epigenetic alterations has received increasing attention for its involvement in various disease processes as well as normal physiological functions. Moreover, it is now generally accepted that DNA methylation and histone modification are imprinted during embryonic development and must be maintained in adults for health and phenotypic stability [12]. Meanwhile, growing evidence suggests that cellular and organismal aging is closely linked to profound changes in a series of epigenetic modifications, leading to altered gene expression patterns [13,14]. In particular, accumulating evidence has demonstrated that epigenetic alterations such as DNA methylation [15–18], histone modifications [19,20], and regulation of ncRNAs [21–23] are involved in the initiation and development of renal aging processes [24]. Recently, aberrant epigenetic modifications, mainly including DNA methylation [11,17], histone modifications [20,25,26], and regulation of ncRNAs [27,28] have also been increasingly implicated in driving age-related kidney disorders. However, no previous systematic review has been performed to systematically generalize the relationship between epigenetics and age-related kidney diseases.

Epigenetics is a rapidly growing field of research and there is a profound interest in exploiting the epigenetics phenomena as diagnostic biomarkers or therapeutic targets of aging-related diseases [29]. In this review, we summarize the current understanding of the pathophysiological roles of epigenetics, particularly focusing on DNA methylation, histone modifications, and ncRNAs modulation, in age-related kidney diseases. At the same time, we focus on the promising use of these epigenetic alterations as potential tools for the early diagnosis, treatment, and prevention of age-related kidney diseases. Taken together, we suggest that epigenetics plays a key role in the aging process and could be used as potential therapeutic targets for the treatment of age-related kidney disease, which warrants further investigation to advance relevant knowledge. 

2. The Phenotype and Mechanism of Renal Aging

2.1. Molecular Mechanisms of Aging 

Aging is associated with an inevitable time-dependent decline in cellular, tissue, and organ functions [30], which has attracted the wide attention and curiosity of researchers throughout the entire history of humankind [6]. Although the free radical theory of aging was proposed as early as in 1956 by Denham Harman [31], it was not until 1983, when the first long-lived strains were isolated from Caenorhabditis elegans, that a new era in aging research began [32]. Currently, the proposed mechanisms that contribute to the aging process include genomic instability, telomere attrition, epigenetic alterations, loss of proteostasis, deregulated nutrient sensing, mitochondrial dysfunction, cellular senescence, and stem cell exhaustion as well as altered intercellular communication. In addition, the aging process is also profoundly influenced by external factors [33] such as bad living habits and adverse environmental factors, which are also associated with the aging of the body [34,35]. In contrast, healthy lifestyle habits such as good eating habits and moderate exercise can effectively delay aging and improve the quality of life of aging populations [34,35]. In spite of the numerous theories that have been proposed during the past four decades to explain the phenomenon of the aging process, the exact mechanisms that drive the fundamental process of aging are not fully understood yet [36,37]

Although the human aging process is associated with a gradual decline in the function of a variety of organ systems, substantial variation exists among people with the same age group due to a combination of genetic and epigenetic factors as well as environmental factors [24]. As such, there is no gold standard for determining what constitutes healthy aging, nor can a single biomarker provide a valid and reliable measure of biological aging [38]. Over the past several decades, a series of high-potential candidate biomarkers of aging have been proposed and evaluated, but none of them have turned out to be universally applicable [39–41]. Clearly, more in-depth research is needed to obtain a systemic understanding of the underlying mechanisms of the aging process to provide an accurate prediction of aging and identify individuals at high risk of developing age-associated diseases or disabilities.

2.2. Pathological Characteristics of Aging Kidney 

Aging is associated with an inevitable time-dependent decline in renal function and increased susceptibility to various acute or chronic kidney diseases [5,6], imposing a huge burden on healthcare systems globally. In order to discover efficient methods to delay renal aging, it is becoming very necessary to identify the factors driving age-associated changes in the kidney [3,42]. The health of the kidney depends on various factors including genetic background, gender, race, oxidative stress, and chronic inflflammation as well as epigenetic factors (e.g., DNA methylation, histone modifications), all of which play a key role in kidney aging process [43]. Although data generated from both cell and animal experiments indicate multiple pathways of potential importance for renal aging (Figure 1), data supporting their involvement in humans are currently rare, with additional research warranted.

Figure 1. Schematic diagram of the main mechanisms and phenotype of renal aging. With increasing age, there are signifificant changes in both the function and structure of the kidney. Multiple age-related pathways contribute to altered renal outcomes in the elderly. 

With increasing age, there are irreversible alterations in the kidney at both microscopic and macroscopic levels as well as clinical and functional. It has been reported that renal weight decreases from the third to eighth decade of life, with the steepest decline occurring after the age of 50 [44,45], but imaging analysis indicates that the kidney parenchymal volume remains unchanged [46]. This may be, in large, attributed to compensatory hypertrophy of unaffected nephrons in response to the loss of nephrons induced by glomerulosclerosis and tubular atrophy [47]. A number of structural changes occur in the kidney with aging, as demonstrated in Figure 1, mainly including a reduced nephron number, decreased total nephron size, glomerular basement membrane (GBM) thickness, glomerulosclerosis, interstitial fibrosis, arteriosclerosis, and tubular atrophy, which ultimately leads to decreased renal plasma fellow (RPF) and glomerular filtration rate (GFR) [3,48]. Moreover, aging kidneys exhibit increased vulnerability to adverse conditions such as oxidative stress, inflflammatory mediators, and fibrotic factors [49,50].

Collectively, these age-related structural and functional changes in the kidney may inevitably predispose the kidneys to various acute or chronic kidney disorders. 

2.3. Age-Related Kidney Dysfunction 

To a better understanding of the aging process, to effectively intervene in it, distinguishing the difference between the natural aging process and age-related dysfunction is especially important [51]. This point is highly pertinent to a variety of age-related diseases but is especially relevant in the context of kidney disease. Although aging itself does not cause kidney disease, age-related structural and functional changes may predispose aging individuals to various kidney diseases [7]. For example, under adverse conditions such as ischemia-reperfusion injury (IRI) and nephrotoxicity insult toxins, elderly people have increased susceptibility to AKI [42,52]. Several studies have demonstrated that the incidence of CKD in the elderly is 3–13 times higher than that in younger individuals [53,54]. Besides, there is evidence that cellular senescence is engaged in the pathogenesis of diabetic kidney disease (DKD), and that hyperglycemia also contributes to cellular senescence in DKD [55,56]. Although renal cellular senescence is the underlying cause of age-related parenchymal glomerular or tubular cell shedding or loss, cellular senescence does not always play an adverse role in all renal diseases. For instance, autosomal dominant polycystic kidney disease (ADPKD) is due to uncontrolled proliferation of renal tubular epithelial cells, and the CDK inhibitor, roscovitine, could mitigate the progression of ADPKD mainly by promoting cellular senescence [57,58]. Currently, although the characteristic alterations of aging kidneys have been well described, the distinction between normal aging and age-related kidney diseases merits further elucidation. Understanding the etiologies of renal aging and age-related renal diseases might enable the rational development of pro-phylactic interventions as well as novel targeted treatment strategies for renal dysfunction. The results of the current studies suggest that epigenetic changes induced by various etiologies (e.g., uremia, cellular senescence, psychological, lifestyle, and various pathogens) may precipitate the development of age-related kidney diseases (Figure 2) [24]. Meanwhile, it is increasingly recognized that epigenetic modifications in embryonic and adult kidney development are interconnected and dysregulation influences age-related kidney diseases [12]. Thus, a further understanding of epigenetic modification regulation in embryonic and adult kidney development will improve our understanding of molecular mechanisms of age-related kidney diseases and develop new treatments against these diseases.

Taken together, from the current studies, we can conclude that despite great progress being made in the understanding of the renal aging process and the associated decline in renal function, the pathogenesis of age-related kidney diseases still remains unanswered and warrants further investigation.

Figure 2. The epigenetic landscape mediates the interplay between etiologies and age-related renal dysfunctions. Both exogenous and endogenous etiologies can lead to changes in the epigenetic landscape that impact renal aging and physiology

3. Epigenetics and Age-Related Kidney Diseases 

Currently, the mechanisms responsible for age-related decline in organ function have not been fully understood, but growing evidence suggests that epigenetic alterations, primarily including aberrant DNA methylation, histone post-translational modifications, and regulation by ncRNAs, play an important role in various age-related human disorders such as neurodegenerative [59], cardiovascular diseases [60], and degenerative spinal stenosis [61] as well as various kidney diseases [24,62]. In the following sections, we will summarize the current knowledge on the implications of epigenetics in age-related kidney diseases. 

3.1. DNA Methylation in Age-related Renal Diseases

Although covalent modifications of DNA bases had been described since 1948 by Hotchkiss [63], it was not until 1969 that Griffifith and Mahler proposed that these modifications may involve the modulation of gene expression [64]. In particular, the major modification in eukaryote DNA is 5-methylcytosine (5mC) [65], which primarily occurs at the fifth position of the pyrimidine ring of cytosines. DNA methylation, the first identified epigenetic mechanism, which occurs primarily at cytosine-phosphate-guanine (CpG) dinucleotides within the gene promoter regions [66], is involved in regulating gene expression through inhibiting specifific transcription factors binding to DNA or recruiting mediators of chromatin remodeling (e.g., histone-modifying enzymes) [67–70]. In mammals, DNA methylation patterns are routinely established and maintained by three DNA methyltransferases (DNMT) [70,71] including DNMT1, DNMT3a, and DNMT3b, while in contrast, DNA demethylation can be achieved by the ten-eleven translocation (TET) enzymes through converting 5mC to 5-hydroxymethylcytosine (5hmC) [72,73].

The dynamic regulation of DNA methylation and demethylation is one of the most important epigenetic regulatory mechanisms in eukaryotic cells, which up to now, has not been fully understood. Increasing evidence has demonstrated that aberrant DNA methylation of specific CpG sites may serve as sensitive biomarkers to identify individuals at risk for age-related diseases [61,74]. In particular, age-related renal diseases such as CKD and ESRD are a major public health problem worldwide because of its high morbidity among aging populations. Several studies have well explored the associations between DNA methylation patterns and age-related kidney diseases [75–77]. For instance, a recent study investigated the genome-wide changes in DNA methylation in renal biopsy samples obtained from 95 healthy kidney donors aged from 16 to 73 years old [11]. A total of 92,778 CpG methylation sites were identified to be significantly associated with donor age through the analysis of genome-wide changes in DNA methylation (more than 800,000 CpG sites) (FDR <0.05), corresponding to 10,285 differentially methylated regions. Interestingly, these regions were most frequently located in the Wnt/β-catenin signaling pathway-related genes including the dickkopf Wnt signaling inhibitors (DKK), several SOX transcription factors, Wnt inhibitory factor 1 (WIF1), secreted frizzled-related protein 2 (SFRP2), retinoic acid receptor alfa and beta (RARA and RARB), and so on. Hypermethylation in the promoter region of these Wnt signaling inhibitor genes may contribute to the activation of Wnt/β-catenin signaling in the aged kidney. Furthermore, Wnt/β-catenin signaling, a conserved signaling pathway in organ development, is kept silent in normal adult kidneys [78,79], which is reactivated predominately in tubular epithelial cells in a variety of CKD models [80]. Thus, hypermethylation of these Wnt signaling inhibitor genes induces activation of Wnt/β-catenin signaling, which may lead to aging-related renal changes by triggering tubular epithelial cell transition to mesenchymal or senescent phenotype and promoting renal fibrosis. This study clearly revealed a causal relationship between DNA hypermethylation and age-associated renal dysfunction [11], indicating that DNA methylation alterations could be a new class of potential non-invasive diagnostic and prognostic biomarkers for age-related kidney diseases. Moreover, numerous clinical observations and animal studies have demonstrated that DNA methylome alterations are implicit in the development and progression of CKD [81–84]. For example, an epigenome-wide association study (EWAS) was performed to investigate the genome-wide methylation profiles in whole blood samples from 4859 aging adults, which demonstrated that the epigenetic signatures were significantly associated with kidney function and CKD as well as with the clinical endpoint renal fibrosis [84]. In this study, the researchers identified 19 CpG sites associated with eGFR and CKD from whole blood samples, among which five CpG sites were associated with renal fibrosis and showed consistent and significant DNA methylation changes in renal cortical biopsy samples from CKD patients. The study revealed that eGFR-associated CpG sites were significantly enriched in regions bound to serval transcription factors including Early B-cell Factor1 (EBF1), E1A Binding Protein P300 (EP300), and CCAAT/enhancer-binding protein beta (CEBPB), highlighting the impact of epigenetic modifications on renal function. Moreover, previous studies have demonstrated that several targeted genes regulated by CEBPB, EBF1, and EP300 are essential for kidney development and function [85–88], suggesting that methylation alterations of CEBPB, EBF1, and EP300 target genes may block the regulation of CEBPB, EBF1, and EP300 on their target genes, leading to the development of CKD. Thus, CEBPB, EBF1, and EP300 may serve as promising candidates for future experimental studies to illuminate the underlying gene regulatory mechanisms linking differential DNA methylation to kidney function in health and disease. 

Despite several genome-wide association studies and epigenome-wide association studies have identified significant changes in DNA methylation with aging and age-related kidney diseases, there is currently still a lack of direct evidence indicating that alterations in particular gene expression patterns as well as gene-specific DNA methylation influence renal aging [17,18]. A more recent study has provided strong evidence for uncovering important epigenetic features of kidney aging [17]. Recently, Gao and colleagues reported that chronic injection of D-galactose (D-gal)-induced aging or natural aging kidneys led to signifificant inhibition of KLOTHO and antiaging factor nuclear factor erythroid-derived 2-like 2 (NRF2) expression, accompanied by increased expression of DNMTs (subtypes of DNMT1, DNMT3a, and DNMT3b) as well as hypermethylation of NRF2 and KLOTHO gene promoter [17]. Administration of DNA-demethylating agent, SGI-1027 and OLP, effectively reduced DNA methylation of the NRF2 and KLOTHO promoter and alleviated D-gal-induced aging-related structural and functional alteration changes in mouse kidneys. Notably, the anti-renal aging effects of SGI-1027 in D-gal-induced aging mice were signifificantly abolished by silencing KLOTHO in vivo. From this study, we can conclude that dysregulation of DNMT1/3a/3b signifificantly contributes to the kidney aging process and epigenetic intervention with DNA-demethylating agents can mitigate renal aging alterations, suggesting that alteration of particular gene expression patterns and genomic DNA methylation can indeed influence the renal aging process. Thus, developing therapeutic strategies aimed at reversing age-associated adverse epigenetic changes will contribute to the development of novel therapeutic interventions that can delay or alleviate renal aging and age-associated kidney disorders

From current research on DNA methylation in age-related kidney diseases (Table 1), we can conclude that DNA methylation might have exerted critical regulatory functions in both normal renal aging and age-related kidney diseases. Nevertheless, the present studies are far from sufficient to elucidate the molecular mechanisms underlying DNA methylation changes in age-related kidney diseases. Furthermore, most of these investigations lack in vivo experimental validation. Therefore, more systematic studies focused on DNA methylation alterations in age-related kidney diseases and clinical applications are required in the future. 

Table 1. DNA methylation in age-related kidney diseases.