Sequencing analysis of the human glucocorticoid receptor (NR3C1) gene in multiple sclerosis patients

https://doi.org/10.1016/j.jns.2016.02.054Get rights and content

Highlights

  • We found no mutations in the coding region of the NR3C1 gene in MS patients.

  • The polymorphisms p.N363S (rs6195), p.N766N (rs6196) and (rs6188) were detected.

  • None of the identified polymorphisms were associated with the severity of disease.

  • None of the identified polymorphisms were associated with response to glucocorticoids.

  • None of the identified polymorphisms were associated with disease subtypes.

Abstract

Various specific human glucocorticoid receptor (NR3C1) gene polymorphisms have been described in multiple sclerosis (MS) patients and correlated with disease progression, susceptibility and aggressiveness. Herein, we investigated the presence of gene alterations in the entire coding region of the NR3C1 in MS patients of variable clinical status (CIS, RRMS and SPMS) and the association(s) of these alterations with severity of disease (EDSS), response to glucocorticoid (GC) treatment and clinical improvement. Sixty Caucasian Greek MS patients were included. Sequencing the coding sequences and intron-exon boundaries of the NR3C1 did not reveal the presence of mutation(s) in any of the MS patients. Three previously described polymorphisms were detected: p.N363S (rs6195), p.N766N (rs6196) and c.1469-16G > T (rs6188). None of the identified alleles/genotypes were found to be associated with the severity of disease, response to glucocorticoids and disease subtypes. Known polymorphism, such as ER22/23EK that has been previously detected in MS patients, was not detected. There is a considerable ethnicity-related variation in the frequency of the NR3C1 polymorphisms. Although a genetic basis of the glucocorticoid sensitivity exists in healthy population, in the presence of chronic inflammation and abundance of cytokines - such in MS patients - other factors appear to play a more important role in GC sensitivity.

Introduction

Accumulating evidence suggests that patients with multiple sclerosis (MS) display increased hypothalamic-pituitary-adrenal (HPA) axis activity, while the lack of their response to the combined Dex-CRH suppression test [5], [25] implicates an impaired negative feedback and a loss of sensitivity to glucocorticoids in the central nervous system. In addition, patients with MS may also demonstrate peripheral glucocorticoid (GC) resistance, as determined by decreased ex vivo dexamethasone inhibition of lipopolysaccharide (LPS)-induced interleukin (IL)-6 or tumor necrosis factor (TNF)-α production in peripheral blood lymphocytes compared with healthy individuals [4]. Notably, central and peripheral GC sensitivity have been associated with clinical MS disease characteristics and clinical improvement [4], [24], [25]. The human glucocorticoid receptor (hGR, hGRα and hGRβ isoforms) is a key regulator of the negative feedback mechanism and of the peripheral GC sensitivity, therefore a defect in the number and/or function of hGR could be a cause for the impaired HPA function and GC resistance in peripheral blood lymphocytes in patients with MS. Various mutations or polymorphism(s) of the hGR gene are reported to alter hGR activity, resulting in enhanced or diminished GC sensitivity [7]. However, no mutation data of the entire hGR (NR3C1) gene in patients with MS have been, so far, reported. Existing studies have investigated various hGR gene polymorphisms in MS patients and correlated these polymorphisms, with disease progression, susceptibility and aggressiveness. van Winsen et al. [23] have demonstrated that among the hGR gene polymorphisms N363S (rs6195), ER22/23EK (rs6189) and the BclI (rs41423247), the ER22/23EK polymorphism is associated with a more aggressive MS phenotype. In a subsequent study, none of the haplotypes of hGR gene (TthIIIΙ (rs10052957), ER22/23EK, BclI, N363S and GR-9β (rs6198) were associated with disease susceptibility, age at onset or onset type [24]; however, MS patients carrying the haplotype 6 (TthIIIΙ, ER2223EK, and GR-9β) have a more rapid disease progression, possibly due to the presence of ER22/23EK and not of GR-9β G and TthIIIΙ polymorphisms [24]. Finally, van Winsen et al. [22] showed that the variants N363S, BclI or ER22/23EK were not associated with GC sensitivity in patients with MS.

To further elucidate the role of hGR gene in MS, we investigated the presence of gene alterations in the entire coding region of hGR in MS patients of variable clinical status, such as clinical isolated syndrome (CIS), relapsing/remitting multiple sclerosis (RRMS) and secondary progressive multiple sclerosis (SPMS). We also investigated the association(s) of alterations in the hGR gene with severity of disease, response to glucocorticoid (GC) treatment and clinical improvement.

Section snippets

Patients

Sixty Caucasian Greek patients fulfilling the revised McDonald criteria for MS or possible MS [13] were included in the study.

Twenty patients (8 males, 12 females) had CIS suggestive of MS, while 22 (4 males, 18 females) had RRMS and 18 patients (9 males, 9 females) had SPMS disability that was assessed through the Expanded Disability Severity Scale (EDSS).

Thirty six out of sixty patients (14/20 CIS, 10/22 RRMS, 12/18 SPMS) were in relapse. These patients received corticosteroids intravenously

Results

Sequencing the coding region of the hGR gene did not reveal the presence of mutation(s) in any of the sixty patients with MS examined. However, three previously described polymorphisms were detected: i) the p.N363S (rs6195), ii) the p.N766N (rs6196) and iii) an intronic variant 16 bp upstream of the exon 5 acceptor splice site, c.1469-16G > T (rs6188).

Testing for heterozygote deficit by application of a score (U) test showed that MS population was in Hardy-Weinberg Equilibrium (p = 0.304).

Discussion

The current study was designed firstly to provide evidence for hGR gene alterations in patients with MS of different disease subtypes, such as CIS, RRMS and SPMS, and secondly to possibly correlate these hGR gene alterations with the severity of disease, response to GC therapy and subsequent clinical improvement. To the best of our knowledge, this is the first time that the entire hGR coding sequence was screened for the presence of mutations and polymorphisms in patients with MS. Utilizing

Declaration of interest

There is no conflict of interest.

Funding

We acknowledge the financial support from the National Kapodistrian University of Athens (Grant number 70/3/9601).

References (25)

  • N.A. Huizenga et al.

    A polymorphism in the glucocorticoid receptor gene may be associated with and increased sensitivity to glucocorticoids in vivo

    J. Clin. Endocrinol. Metab.

    (1998)
  • A.K. Kemppinen et al.

    Systematic review of genome-wide expression studies in multiple sclerosis

    BMJ Open.

    (2011)
  • Cited by (7)

    View all citing articles on Scopus
    View full text