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Author: Tracy
Atrial cardiomyopathy (ACM) is the main culprit of atrial fibrillation (AF) and thromboembolic events. Diabetes mellitus (DM) is an important risk factor for ACM. However, the underlying mechanism between ACM and DM remains elusive.
2024年8月3日， 上海交通大学医学院附属新华医院王群山团队在期刊《eBioMedicine》上发表了题为“NR4A3 prevents diabetes induced atrial cardiomyopathy by maintaining mitochondrial energy metabolism and reducing oxidative stress”的研究论文。 本研究首次证实了糖尿病心房中NR4A3的减少，直接导致SDHA的转录下调。 本研究揭示了NR4A3在糖尿病诱导的ACM中的关键作用，为心房肌病提供了新的潜在治疗靶点。
https://www.thelancet.com/journals/ebiom/article/PIIS2352-3964(24)00304-9/fulltext#%20
Background:
01
Atrial cardiomyopathy (ACM) is of increasing clinical and research interest and can be seen as a precursor state of atrial fibrillation (AF) and stroke. AF itself can be seen as electrical remodeling in ACM, which can also exacerbate ACM by inducing structural and functional changes in the atria, such as myocardial hypertrophy, ion channel dysfunction, and loss of rhythm control. Therefore, AF and ACM have a complementary and inseparable relationship.
Diabetes, hypertension, older age, and heart failure can worsen ACM. Diabetes mellitus is one of the most prevalent chronic diseases and an independent risk factor for AF. Patients with diabetes have an approximately 40% increased risk of developing atrial fibrillation, while patients with atrial fibrillation and diabetes have significantly higher rates of hospitalization, cardiovascular mortality, and all-cause mortality. Diabetes mellitus can enhance susceptibility to atrial fibrillation through a variety of mechanisms, including atrial structural remodeling, electrical remodeling, electromechanical remodeling, autonomic remodeling, inflammation, and oxidative stress.
Excitation-contraction coupling and electrical conduction in atrial cardiomyocytes require massive adenosine triphosphate (ATP) support. Disorders in mitochondrial energy production in atrial myocytes can lead to atrial remodeling, which contributes to the development of arrhythmias, including atrial fibrillation. Decreased expression of multiple tricarboxylic acid (TCA) cycle key enzymes (e.g., aconitate hydratase, citrate synthase, and malate dehydrogenase) was observed in patients with AF compared to patients with SR. In addition, dysfunction of mitochondrial electron transport chain (ETC) complexes I and II was observed not only in isolated patients with AF, but also in patients with diabetes mellitus. The scavenging of reactive oxygen species (ROS) relies on the involvement of various key enzymes in the TCA cycle, such as nicotinamide adenine dinucleotide (NADH) and nicotinamide adenine dinucleotide phosphate (NADPH). As a result, the reduction in TCA cycle turnover leads to a significant increase in ROS production, which further promotes atrial remodeling. It is also clear that AF is associated with increased oxidative stress.
The NR4A subfamily (nuclear receptors of subfamily 4) belongs to the nuclear receptor family and, along with members such as NR4A1 (also known as NUR77), NR4A2 (NURR1), and NR4A3 (NOR1), acts as transcription factors that can respond rapidly to various stimuli, such as growth factors and hormones. All three receptors share a common structure, including a ligand-independent activation function (AF)-1 transactivation domain, a ligand-dependent AF-2 transactivation domain, and a DNA-binding domain (DBD). The DBD domains of these receptors exhibit 90%–95% homology and can bind to NBRE (nerve growth factor [NGFI-B] response element), which consists of octamer sequence (consensus sequence AAAGGTCA) zeros, or two inverted NBREs, consisting of a limited number of nucleotides, as homodimers or heterodimers. The high degree of homology of DBDs ensures functional consistency within the NR4A family, although their activity is also affected by transcriptional levels and post-transcriptional modifications. Overexpression of NR4A3 in muscle can enhance the intermediate content of TCA circulation and electron respiratory chain conduction, including isocitrate dehydrogenase 3 (IDH3), fumarate dehydrogenase (FH), and ATP synthesis F1 α (ATP5a1).
In this study, the team revealed for the first time that NR4A3 is highly expressed in the atria and has a protective effect on diabetes-induced ACM. The team found a significant reduction in NR4A3 in atrial tissue samples from AF patients and diabetic mice. Mechanistically, NR4A3 deficiency promotes mitochondrial energy metabolism dysfunction and oxidative stress damage by inhibiting SDHA transcription levels, thereby aggravating atrial remodeling.
Research Progress
02
NR4A3 inhibits diabetes-induced atrial structural remodeling
The experimental results showed that compared with db/m mice, db/db mice had significant hepatic steatosis, significantly enlarged adipocytes, and increased serum metabolic indexes ALT, AST, TG and TC. In addition, glucose tolerance and insulin resistance are markedly impaired. However, overexpression of NR4A3 in db/m mice or db/db mice showed no significant change in the above phenotype.
Overexpression of NR4A3 in db/m mice does not produce any significant changes in LAW/TBW ratio, left atrial area, atrial fibrosis, and atrial cardiomyocyte cross-sectional area in db/m mice compared to ctrl db/m mice. In contrast, upregulation of NR4A3 in db/db mice significantly attenuated the LAW/TBW ratio, left atrial area, atrial fibrosis, and atrial myocyte cross-sectional area compared to db/db mice bearing AAV9-Ctrl. In db/m mice, overexpression of NR4A3 had no significant effect on ANP, COL1A1 and COL3A1. However, in db/db mice, overexpression of NR4A3 resulted in a significant decrease in ANP, COL1A1, and COL3A1 expression. Therefore, overexpression of NR4A3 can alleviate atrial fibrosis and cardiomyocyte hypertrophy in db/db mice.
IL-6 and TNF-α levels were mildly elevated in both diabetes models. However, overexpression of NR4A3 reduced the levels of IL-6 and TNF-α in db/db mice, while knockdown of NR4A3 exacerbated the levels of IL-6 and TNF-α in the atrial tissues of HFD+STZ-treated T2DM mice. There was no significant change in the expression of IL-1β in the diabetes model. The results showed that the overexpression of NR4A3 could reduce inflammation in diabetic mice.
Figure 3: Heart-specific overexpression of NR4A3 attenuates diabetes-induced atrial remodeling. (A) Representative whole-heart images of db/m or db/db mice at 16 weeks after injection of AAV9-cTNT-Nr4a3 or AAV9-cTNT-Ctrl (scale bar=1 mm). (B) Representative image of the long axis of a two-dimensional echocardiogram. (C) Representative image of Masson staining (scale bar = 50 μm) (D) Representative image of WGA staining (scale bar = 20 μm). (E) Ratio of left atrial weight (LAW) to total body weight (TBW) (LAW/TBW, n=8 in each group). (F) Left atrial area in each group (n=8 in each group). (G) Quantitative results for fibrosis area in the specified groups (n = 8 per group). (H) Quantitative results of cardiomyocyte area in indicator groups (n=8 in each group). (I and J) immunoblotting (I) and quantification (J) of ANP protein levels (n=6 per group). (km) immunoblotting (K) and quantification (L and M) of COL1A1 and COL3A1 protein levels (n=6 per group). All data are presented as standard deviations ± averages. For statistical analysis, one-way ANOVA versus Bonferroni post-hoc analysis was used.
SDHA过表达减弱糖尿病Nr4a3-KO小鼠的心房重塑
Given the critical role of SDHA in NR4A3-mediated diabetes-induced ACM regulation, Nr4a3-KO mice receive injections of SDHA-expressing AAV9 vehicle or control at 8 weeks followed by 16 weeks of treatment with ND or HFD+STZ. The results suggest that upregulation of SDHA inhibits NR4A3 deficiency-induced LAW/TBW, left atrial size, atrial fibrosis, remodeling of the cross-sectional area of atrial cardiomyocytes, and susceptibility to pacing-induced AF in diabetic mice. Transmission electron microscopy showed that the overexpression of SDHA did not significantly change the size and number of mitochondria in diabetic Nr4a3-KO mice. However, it significantly alleviates mitochondrial damage while significantly improving ATP production.
Figure 8: Atrial remodeling attenuated by SDHA overexpression in diabetic Nr4a3-/- mice. (A) Representative whole-heart images of Nr4a3-/- mice injected with AAV9-SDHA or AAV9-CTRL 16 weeks after ND or HFD+STZ treatment. (B) Representative image of the long axis of a two-dimensional echocardiogram. (C) Representative image of Masson staining (scale bar = 50 μm). (D) Representative image of WGA staining (scale bar = 20 μm). (E) Ratio of left atrial weight (LAW) to total body weight (TBW) (LAW/TBW, n=6 in each group). (F) Left atrial area in each group (n=6 in each group). (G) Quantitative results for fibrosis area in the specified groups (n= 6 per group). (H) Quantitative results of cardiomyocyte area in the indicator group (n=6 in each group). (I) Representative ECG trajectories (n=6 per group). (J) Probability of successful induction rate in a total of 10 burst stimuli for each group. (K) Duration of AF in the indicated group. (From left to right) transmission electron microscopy (TEM) images (L) and quantitative results of mitochondrial size (M), number of mitochondria/μm 2 (N) and mitochondrial vacuolar ratio (O) (scale bar = 1 μm and 500 nm). (P) ATP production (n=6 per group).
Conclusions of the study
03
NR4A3 acts as a transcription factor that regulates the transcription of various disease-related genes. However, there have been no previous reports of NR4A3 involvement in SDHA regulation. In this study, the team presented a new discovery that NR4A3 does regulate SDHA transcription. The team elucidated a specific NR4A3 response element in the SDHA promoter region through a Cut&Tag assay and a dual luciferase reporter assay. In addition, the overexpression of SDHA in Nr4a3-/- diabetic mice showed a remissional effect on atrial remodeling, providing valuable insights into the potential therapeutic role of NR4A3 in diabetic atrial pathology.
There are interactions between the various organs in the body, and a variety of factors can contribute to multi-organ dysfunction. For example, metabolic risk factors such as obesity, insulin resistance, abnormal blood glucose levels, and dyslipidemia often cluster together and are major risk factors for cardiovascular disease (CVD), T2DM, and NAFLD.
Several studies have confirmed that NAFLD and obesity increase the risk of developing AF. They can cause metabolic disorders, including increased insulin resistance and cardiac lipidosis, which can lead to atrial remodeling. In addition, by releasing inflammatory cytokines and increasing oxidative stress, NAFLD and obesity can also lead to atrial fibrosis, LA enlargement, electrical remodeling, and autonomic remodeling. NAFLD and obesity cause an increase in the volume and thickness of epicardial adipose tissue, releasing adipokines that act on the atrial tissue through paracrine effects, leading to atrial remodeling.
The team used the AAV9 vector with a cTNT promoter to overexpress NR4A3. Subsequently, the team assessed the morphology and function of the islets, liver, and adipose tissue. The results confirmed that the effects of NR4A3 overexpression were concentrated only in the heart. Therefore, the protective effect of NR4A3 on diabetes-induced atrial remodeling stems from its direct effect on the heart itself.
However, in Nr4a3-KO mice, the team was unable to completely eliminate potential interactions between systems. Nonetheless, the overall findings confirm that NR4A3 deficiency exacerbates diabetes-induced atrial dysfunction.
Resources:
1.Goette A. Kalman J.M. Aguinaga L. et al.
EHRA/HRS/APHRS/SOLAECE expert consensus on atrial cardiomyopathies: definition, characterization, and clinical implication.
Europace. 2016; 18: 1455-1490
2.Goldberger J.J. Arora R. Green D. et al.
Evaluating the atrial myopathy underlying atrial fibrillation: identifying the arrhythmogenic and thrombogenic substrate.
Circulation. 2015; 132: 278-291
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