1. Introduction
Amyotrophic Lateral Sclerosis (ALS) is a progressive neurodegenerative disorder that mainly affects the upper and lower motor neurons, while about half of the patients present cognitive decline during the course of the disease [
1]. The worldwide prevalence of ALS is estimated approximately between 4 and 5 patients per 100,000 individuals, whereas its incidence corresponds to about 1 to 2 new cases per 100,000 person-years [
2]. ALS is more common among males and its prevalence follows an upward trend towards the 7th and 8th decades of life [
2]. The mean survival of ALS patients is estimated between 2 and 4 years for most populations [
2]. The only FDA-approved treatments; riluzole and edaravone, provide only small benefits regarding the median survival and clinical progress of the disease [
3,
4].
ALS pathology is characterized by degeneration of motor neurons in the cortex, brainstem motor nuclei and spinal anterior horns [
5]. Multiple mechanisms have been associated with potential motor neurotoxicity in ALS; oxidative stress, mitochondrial dysfunction, glutamate-induced excitotoxicity, protein misfolding and aggregation, impaired axonal transportation and microglia-related mechanisms have been incriminated, but a definite conclusion for the underlying pathophysiology of ALS has not been reached [
6]. Genetic and environmental parameters, as well as genetic-environmental interactions, are considered to contribute to the overall risk of the disease [
7]. About 5–10% of the ALS cases are estimated to be of familial incidence, while the rest of the cases are sporadic [
8]. The most important mutations associated with the familial form of the disease are related to the
C9orf72 and
SOD1 genes, as well as the FUS/TLS and TDP43 RNA binding proteins [
6]. Genetic factors are considered to play an important role in the incidence of sporadic ALS too [
5]. One previous Genome-wide association study (GWAS) has specifically revealed (among others) an association with a new genetic locus,
MOBP (Myelin-associated Oligodendrocyte Basic Protein) at 3p22.1, and particularly the
rs616147 Single Nucleotide Polymorphism (SNP) [
9].
MOBP, like myelin basic protein (MBP), is produced by oligodendrocytes and is located in the major dense line of Central Nervous System (CNS) myelin [
10]. Although it is hypothesized that
MOBP contributes to the compacting and stabilization of the myelin sheath through
MOBP-MBP interactions, its definite function remains unclear [
10]. Oligodendrocytes and myelination processes have a crucial role in several neurodegenerative diseases such as Multiple Sclerosis (MS) [
11] and Alzheimer’ s Disease (AD) [
12], while
MOBP in particular has been associated with both of the aforementioned entities [
13,
14,
15]. Furthermore, there is a relationship between Single Nucleotide Polymorphisms (SNPs) in the
MOBP genetic locus and frontotemporal dementia (FTD) [
16], a disease strongly related to ALS [
1], as well as progressive supranuclear palsy (PSP), an entity of the frontotemporal lobar degeneration (FTLD) spectrum pathology [
17].
Oligodendrocytes and myelination are considered important in the pathogenesis of ALS, as well. This argument is supported by changes in the composition of myelin (even demyelination) [
18,
19] and relevant pathological findings (including dysfunction, degeneration, defective regeneration) in grey matter oligodendrocytes of ALS subjects [
20,
21]. Given this background,
MOBP could be potentially implicated in the pathogenesis of ALS. To date, only the above mentioned GWAS has identified
MOBP, and the
rs616147 SNP (an intron variant -adenosine-guanosine replacement- of the
MOBP gene), as a potentially ALS-associated locus [
9]. Therefore, a case-control study was performed to assess the replicability of the association between
rs616147 and sporadic ALS in patients of Greek ethnicity.
3. Results
All of the patients and controls that were invited to participate in the study responded positively and completed the study processes. A total of 155 patients with definite sporadic ALS and an equal number of age (±2 years) and sex matched healthy controls were recruited. As noted above, the genotype call rate was 99.03%, with a total of 152 ALS patients and 155 healthy controls ultimately having available genetic data. The power of our sample size was slightly above 80% to find a significant association (
p < 0.05) between
MOBP rs616147 and ALS, given a minor (A) allele frequency of 32% [
32], a disease prevalence of 5/100,000 [
2], and an estimated relative risk of 1.60. Patient characteristics are presented in
Table 1.
MOBP rs616147 was determined to be in HWE among healthy participants (
p = 1.00). Allelic and genotypic frequencies are provided in
Table 2. The minor allele (A) frequencies were 29% and 33% for the cases and controls, respectively.
According to the univariate analysis, there was no significant relationship between
MOBP rs616147 and ALS (primary outcome measure) with respect to every mode of inheritance; log-additive OR = 0.85 (0.61, 1.19), over-dominant OR = 0.73 (0.46, 1.15), recessive OR = 1.02 (0.50, 2.09), dominant OR = 0.74 (0.47, 1.16), co-dominant OR
1 = 0.71 (0.44, 1.14) and co-dominant OR
2 = 0.88 (0.42, 1.84) (
Table 3).
Additionally, the effect of
rs616147 on the age of ALS onset was investigated (
Table 4). Both crude (G/G vs. G/A; HR = 1.12 (0.80, 1.59), G/G vs. A/A; HR = 0.91 (0.54, 1.54)) and sex-adjusted (G/G vs. G/A; HR = 1.11 (0.79, 1.56), G/G vs. A/A; HR = 0.94 (0.55, 1.60)) cox-proportional hazards models provided evidence indicative of no association between
rs616147 and age of ALS onset (the effect of sex on the age of ALS onset was determined insignificant; HR = 1.37 (0.96, 1.84)). Subgroup analyses based on the site of onset, reproduced the insignificant associations, both when unadjusted and adjusted for sex (the effect of sex on the age of ALS onset was determined insignificant regarding the limb onset ALS; HR; 1.44, (0.85, 2.44), but significant regarding the bulbar onset ALS; HR = 3.43 (1.29, 9.09), with male sex presenting later onset of the disease) (
Table 5).
Finally, no association was found between
rs616147 and site of ALS onset (
Table 6). Limb onset (vs. bulbar and mixed onset) and bulbar onset (vs. limb and mixed) were separately analysed.
4. Discussion
The present case-control study investigated the effect of
MOBP rs616147 on the development of ALS, as well as the age and site of ALS onset. Results were compatible with the absence of an underlying connection, regarding all outcome measures. Our findings come in contradiction with the findings of the only other study that examined this association, a previous very large GWAS [
9]. The aforementioned study involved data from a great number of ALS cohorts originating from Western countries, but a Greek cohort was not included. Apart from the
MOBP rs616147—ALS direct association, a dose-dependent pharmacogenetic interaction has been recently revealed between the A allele of
rs616147 and creatine administration to patients with ALS [
33].
The functions of
MOBP are not yet completely clarified (the functional significance of the
rs616147 polymorphism as well), but it appears to participate in myelin compaction and stabilization, through interactions with a structurally similar protein, MBP [
10,
34,
35]. Oligodengroglial dysfunction and myelination disorders are crucial in ALS [
36]. Grey matter oligodendrocytes present substantial degeneration in ALS animal models, while precursor cells fail to differentiate and compensate for the losses, ultimately leading to incomplete recovery. The degeneration of oligodendrocytes subsequently causes myelin abnormalities, such as immature myelin sheaths and demyelinated axons, and, finally, leads to axonal degeneration. These pathological findings support the notion that myelination processes (as well as myelination-related loci such as
MOBP) are potentially important for ALS. On the other hand,
rs616147, which is an intron variant of the
MOBP gene, may not directly affect the function of
MOBP and by extension myelination, rendering the myelination abnormalities among ALS patients irrelevant.
The association of
MOBP with other neurodegenerative diseases is well-established. To date, autoreactivity against
MOBP has been detected among individuals with MS [
13,
15] and
MOBP immunoreactivity has been detected in the core of Lewy Bodies (LBs) among patients with Parkinson’ s disease and dementia with LBs [
37,
38].
MOBP SNPs have been associated with Apolipoprotein-E e4 positive AD [
14], FTD (and the severity of white matter degeneration [
16,
39]), PSP [
17,
40,
41,
42,
43,
44], Corticobasal Degeneration [
43,
44], while decreased expression of
MOBP was revealed in familial Globular Glial Tauopathy [
45] and differential DNA methylation of
MOBP was shown in Multiple System Atrophy [
46]. The implication of
MOBP in all these neurodegenerative disorders makes it possible that it is also involved in the pathophysiology of ALS.
The study, nevertheless, failed to demonstrate the existence of an association. The present study was the first to examine this SNP among individuals of Greek ethnicity with sporadic ALS. Therefore, the replicability of our results cannot be tested. At this stage, it is appropriate to recognize that our study has some limitations, which may be accountable for the non-significant results. First, the power of our study was slightly over 80%, which means that there is an almost 20% possibility that we failed to detect a truly significant association. Additionally, both primary and secondary analyses were not adjusted for several potential protective or risk-conferring factors, including both genetic and environmental ones (such as pesticide and metal exposure) [
8,
47]. Therefore, our results may be significantly affected by the latent effect of an uncontrolled parameter. Moreover, both patients and healthy controls (matched for sex and age ±2 years, selected from the same community) were recruited from a specific geographical area located in Central Greece. The selection of this sample achieved several advantages by possibly matching for several undetermined exposures to a variety of environmental factors (e.g., soil and water metal concentrations) but might have induced an overmatching bias when it comes to genetic parameters. Finally, the conduction of a GWAS is more appropriate than the investigation of a single locus for distinguishing disease vulnerability genes in sporadic maladies. Thus, ideally, a large-scale GWAS with Greek ALS patients should be conducted to evaluate the association of
rs616147 with ALS and individuals of Greek ethnicity.