NBR and GBA Gene Methylation Levels in the Peripheral Blood of Parkinson’s Disease Patients

Article information

J Mov Disord. 2024;17(4):456-458

Publication date (electronic) : 2024 July 22

doi : https://doi.org/10.14802/jmd.24111

Yagmur Inalkac Gemici^1,2,, Ahmet Koc¹

¹Department of Medical Biology and Genetics, Inonu University, School of Medicine, Malatya, Turkey

²Department of Neurology, Manisa Celal Bayar University, School of Medicine, Manisa, Turkey

Corresponding author: Yagmur Inalkac Gemici, MD, PhD Departmant of Neurology, Manisa Celal Bayar University, School of Medicine, Uncubozköy mah. 5524. Sok No: 11 B blok kat: 4 daire: 13 Yunusemre, Manisa, Turkey / Tel: +90-5432517710 / E-mail: yagmur.gemici@cbu.edu.tr

Received 2024 May 9; Revised 2024 June 13; Accepted 2024 July 19.

Dear Editor,

Parkinson’s disease (PD) is a neurodegenerative disorder characterized by the accumulation of Lewy bodies (LBs) in the substantia nigra. Properly functioning protein quality control systems, such as the unfolded protein response and autophagy, are crucial for neuronal health. Autophagy dysfunction is primarily implicated in neurodegenerative diseases such as Alzheimer’s disease, PD, Huntington’s disease, multiple sclerosis, and amyotrophic lateral sclerosis [1]. Autophagy is a well-established pathophysiological mechanism in PD development. While many studies have focused on abnormalities in the central nervous system, recent research has identified peripheral and enteric nervous system anomalies [2]. We hypothesized that changes related to autophagy could also be detected in the peripheral blood of PD patients.

To evaluate autophagy in PD, we aimed to assess a gene known to be significant in PD and associated with autophagy, along with a gene that serves as a primary marker of autophagy but has not been identified in PD patients. Therefore, we selected one of the following genes, LRRK2, DJ-1, PINK1, Parkin, or GBA, as well as the NBR1 gene, which encodes the primary receptor protein for autophagy but has not been identified in PD.

Our cross-sectional prospective study was conducted at the Departments of Neurology and Medical Genetics at Inonu University, Malatya, Turkey. We included patients who met the UK PD Society Brain Bank clinical diagnostic criteria [3]. Clinical findings were assessed via the Unified Parkinson’s Disease Rating Scale (UPDRS). The exclusion criteria for the study included the presence of diseases associated with autophagy, such as diabetes mellitus, cardiovascular diseases, cancer, fatty liver disease, autoimmune diseases, and kidney disease. Additionally, individuals on PD-related medications or those with an active infection at the time of blood collection were excluded.

DNA was extracted from the peripheral blood of PD patients (n = 20) and controls (n = 20). The extracted DNA samples were digested with methylation-sensitive restriction enzymes (HpaII and Hin6I; Fermentas, Vilnius, Lithuania) for 24 hours, and the digestion efficiency was confirmed by running the samples on 1% agarose gels. We used a methylation-specific quantitative polymerase chain reaction approach to assess the methylation status of the genes. Gene primers (Supplementary Table 1 in the online-only Data Supplement) and protocols are provided in the Supplementary Material (in the online-only Data Supplement).

The mean age of the PD patients was 62.05 ± 7.03 years, while the mean age of the controls was 58.15 ± 4.35 years (p = 0.102). The supplementary material includes a dot plot and a statistical analysis of participants before the removal of outliers (Supplementary Figure 1 and Supplementary Table 2 in the online-only Data Supplement), which revealed outliers in terms of gene methylation levels. After these outliers were excluded, the statistical analyses were repeated, and the results are presented in Table 1. The significance of age and sex remained unchanged following the removal of outliers (after removal of NBR1 gene methylation participants: PD Group n = 20, control n = 13, age significance p = 0.102; sex [F/M] for PD: 9/11, for control: 5/8, significance p = 0.497; for GBA: PD n = 18, control n = 13, age significance p = 0.196; sex [F/M] for PD: 8/10, for control: 5/8, significance p = 0.394). The methylation levels of NBR1 and GBA were lower in PD patients than in controls (p = 0.005 and p = 0.028, respectively). No correlation was found between the methylation levels of the genes and age or UPDRS score. According to Table 1, the NBR1 and GBA genes, particularly the NBR1 gene, were significantly hypomethylated in PD patients compared with those in controls. These findings suggest that the indirect expression of the autophagy-related genes NBR1 and GBA is elevated in the peripheral blood of PD patients.

Table 1.

Clinical charactaristics and gene methylation levels of participants, compared to controls

NBR1 has been shown to have a neuroprotective effect by enabling LB aggregates to enter the autophagosome in synucleinopathies [1]. However, there are no data in the literature concerning epigenetic regulation affecting NBR1 gene expression in PD patients. In a mouse model of LB disease, the protein and mRNA levels of NBR1 were increased compared with those in controls [4]. Our study demonstrated that autophagic pathways are upregulated by decreased methylation levels of NBR1 in the peripheral blood of PD patients and that autophagy is active in the peripheral system of PD patients, similar to the central system. Further studies should investigate NBR1-related molecules as potential biomarkers in the peripheral system of PD patients.

GBA is one of the genes most significantly associated with PD. Gaucher disease, a lysosomal storage disease, is caused by a deficiency in the enzyme GBA, which interacts bidirectionally with alpha-synuclein to generate positive feedback. Alpha-synuclein aggregation is increased due to GBA mutations [5]. None of the PD patients in our study had a history of Gaucher disease. One study reported greater GBA enzyme activity in PD patients without GBA mutations than in those with mutations and in controls [6]. To the best of our knowledge, there are no data on GBA methylation levels in PD patients. In our study, the GBA methylation level was also decreased in PD patients, similar to NBR1 methylation. GBA-related molecules could be useful biomarkers in the peripheral blood.

The role of DNA methylation in neurodegenerative diseases is an area of intense research. Studies suggest that DNA methylation may contribute to the pathogenesis of PD [7]. In our study, we selected genes relevant to the pathophysiology of PD and found significant differences between the groups. However, no correlation was found between gene methylation levels and age or the UPDRS.

Recent studies on DNA methylation in the blood cells of PD patients have highlighted significant epigenetic changes that may serve as potential biomarkers for early detection and understanding of the disease [7]. Additionally, the integration of gene expression and DNA methylation data revealed dysregulation of genes associated with neurotransmission and protein degradation pathways, suggesting that DNA methylation patterns could reflect environmental exposures and genetic predispositions, offering a new approach for PD diagnosis and progression monitoring [8].

In conclusion, the methylation levels of the NBR1 and GBA genes are lower in the peripheral blood of PD patients than in that of controls. These findings suggest that autophagy-related changes are also present peripherally. NBR1 and GBA-related molecules should be investigated in future studies as potential PD biomarkers.