Background
Apolipoprotein E (apoE) exerts pleiotropic biological functions, including effects on lipoprotein metabolism as well as on the innate and adaptive immune system. Potential mechanisms underlying the immunomodulatory properties of apoE involve enhanced anti-inflammatory macrophage phenotype, decreased activation of NF-kB and STAT1 [
1], and downregulation of TH-1 and TH-17 responses via suppression of pro-inflammatory cytokines secreted by macrophages [
2]. apoE is expressed in the CNS and is produced by antigen-presenting cells (dendritic cells, macrophages). These observations have led to the investigation of
apoE in multiple sclerosis (MS) and its animal model experimental autoimmune encephalomyelitis (EAE; reviewed by [
3]). In this context, controversial results have been reported for
apoE in EAE including both beneficial as well as aggravating effects on disease severity and progression in
apoE knock-out mice [
4,
5]. In parallel, two
APOE polymorphisms, i.e., rs429358 (ε4, Cys130Arg) and rs7412 (ε2, Arg176Cys), which represent established risk variants in Alzheimer’s disease [
6], have been assessed extensively for their role in MS. A recent study compiling data on nearly 30,000 subjects showed that these polymorphisms do not influence MS susceptibility [
7]. However, their role in disease progression still remains ambiguous, which at least in part pertains to the fact that the majority of studies have assessed rather small, i.e., underpowered datasets (for an overview see e.g., [
8]). Divergent findings may also be due to confounders or effect modifiers such as sex, age, or patient subgroups. Along these lines, a comparatively small study testing 221 patients suggested that the association between
APOE and MS severity was limited to women [
9]; however, this has not been described in other studies [
8]. Thus, in the current study, we comprehensively assessed the role of apolipoprotein E on disease severity of EAE as well as MS by taking into consideration potential sex-specific effects of
APOE genotypes.
Discussion
Results of this study indicate that the absence of apoE slightly attenuates EAE in male mice but at the same time aggravates disease course in female animals. In line with this observation, decreased NfH concentration in male apoE-deficient mice in comparison to wt mice suggests an attenuation of axonal damage in male mice lacking apoE. Increased apoE expression in the spinal cord of female and male wt mice in the chronic disease phase may indicate an influence of apoE on disease progression during EAE.
In contrast to the results in the rodent model, we did not detect a robust association between MSSS and APOE rs7412 or rs429358 in over 3200 patients despite excellent (>90 %) power to observe even moderate changes in the MSSS. This suggests that rs7412 and rs429358 do not have a notable influence on MS severity.
Studies that investigated the role of
apoE deficiency in EAE have yielded inconsistent, in parts, and even contradictory results [
2,
4,
5,
23]. Discrepancies may be due to methodological differences (e.g., the immunization protocol) or due to other modifying factors that have not been investigated in the respective studies. In this context, one potential influencing factor that was not controlled for in previous studies is the sex distribution of the tested animals.
apoE is expressed in the CNS in resident immune cells and has been implicated in different immunoregulatory functions [
1,
2,
23]. For instance, in a recent study, milder disease in
apoE-deficient mice was associated with a reduction of dendritic cells (DCs), which—in turn—can be modulated by sex hormones, i.e., estrogens and primarily E2 [
24,
25]. Other studies have reported that apoE modulates macrophages toward an anti-inflammatory phenotype [
1] and suppresses microglial activation [
26,
27]. The activity of these cells can be modulated by the exposition of estrogen and testosterone (reviewed in [
28]). Androgene-receptors (AR) are expressed on immune cells [
29]; therefore, especially, androgens may have immunomodulatory or even immunosuppressive effects [
28]. A direct interaction of apoE with AR has also been described [
30‐
32]. Thus, immune functions appear to be influenced by androgens via AR and may be modulated by apoE. Although we did not investigate mechanistic pathways, the previously described interactions between immune functions and sex hormones may account for some of the sex-specific differences observed in our study that may additionally be influenced by apoE.
While our human data do not reveal sex-specific association of
APOE genotypes and MS severity, the association of
APOE genotypes with Alzheimer’s disease (AD) has been described to be modulated by sex and ethnicity. Whereas
APOE2 and
APOE3 seem to be protective across ethnic groups,
APOE4 increases AD risk [
6]. The latter effect appears to be pronounced in women [
33].
The lack of association of tested
APOE polymorphisms with MS severity is in line with the results of most previous publications (for an overview see [
8]), including a large pooled re-analysis of previously published datasets on 3518 patients [
34] that are independent from those analyzed here. Overall, the authors of the latter study did not find compelling evidence for an association of
APOE and MSSS either. While they observed a higher MSSS in male homozygote carriers of the
APOE e4 allele when compared to all other groups (
p = 0.004), this finding did not withstand multiple comparison corrections [
34]. In light of the fact that the two largest, independent, and well-powered studies on rs7412 and rs42935 did not produce robust results, it appears most likely that rs7412 and rs429358 in
APOE do not play a substantial role in MS severity as measured by the MSSS. However, several aspects need to be considered upon interpretation of our association results: We have tested two
APOE variants, namely two non-synonymous polymorphisms that represent the most important contributors to Alzheimer’s disease risk [
6] and that have been extensively characterized functionally. However, even homozygosity at either of these polymorphic sites does not fully mimic the rodent
apoE knock-out model [
35]. Therefore, the lack of robust genetic effects in humans does not necessarily contradict the results obtained in the
apoE
−/−
mouse model. However, the translation of findings from experimental models and especially in the context of EAE to the human situation has repeatedly failed as only certain facets of the human disease can be modeled [
36]. Thus, we cannot exclude that the effects of apoE observed in the rodent EAE model in this study are of lesser or no relevance for the human disease. In addition, we have only assessed the aforementioned two non-synonymous polymorphisms in the
APOE region. Thus, we cannot exclude the presence of other variants in the
APOE locus with a possible effect on MS severity, although a recent genome-wide association study (which did not assess those two variants directly due to technical reasons (see [
7] for explanation)), did not observe evidence for an association of MSSS and other genetic variants in the
APOE region [
37]. Another consideration extends to the fact that genetic association analyses of MS severity have overall only yielded rather limited success [
38]; one explanation, which could also affect the MSSS association analysis results presented here, is the lack of more appropriate clinical and paraclinical classification schemes to better represent disease severity and progression. In addition, other variables, e.g., information on treatment regimes, may represent confounders in the
APOE association analysis that could not be accounted for in our and previous analyses (e.g., [
34]).
Acknowledgements
We are grateful to the patients and control individuals participating in this study. We thank Ms. Brit-Maren Schjeide for excellent technical assistance. We thank ICM, Généthon for their help and support. French DNA samples were provided by the BRC-REFGENSEP (BB-0033-00019). We thank K. Hofmann for expert technical assistance.
Disclosure of all authors and co-authors
Study sponsorship or funding: This project was funded by grants from the Cure Alzheimer's Fund (to L.B.) and the German Ministry for Education and Research (BMBF; grant 16SV5538 to L.B., KKNMS to F.Z., grant NBL3 to U.K.Z.). The research leading to these results has received funding from INSERM, AFM and the program “investissements d'avenir” ANR-10-IAIHU-06.
A.S. received personal compensation for activities with Novartis, Sanofi and Almirall Hermal GmbH.
L.A.G has received personal compensations (travel support) from Novartis, Biogen Idec, Genzyme/Sanofi Aventis, Teva, Merck Serono, and Bayer Schering Pharma.
F.Z. has received research grants from Teva, Merck Serono, Novartis and Bayer as well as consultation funds from Teva, Merck Serono, Novartis, Bayer Healthcare, Biogen Idec Germany, ONO, Genzyme and Sanofi Aventis. Her travel compensation has been provided for by the aforementioned companies.
T.K. has received travel expenses and personal compensations from Bayer Healthcare, Teva Pharma, Merck Serono, Novartis, Sanofi Aventis and Biogen Idec as well as grant support from Bayer Schering AG and Novartis.
M.B. received a travel grant from Biogen Idec, travel grants, speaker and consultancy honorarium, and a research grant from Merck Serono GmbH, served on an advisory board and received a travel grant from Almirall Hermal GmbH, and received a travel grant and research grants from Teva GmbH as well as from Novartis Pharma GmbH.
U.Z. received personal compensation and research support from Almirall, Bayer, Biogen Idec, Merck Serono, Novartis, Sanofi Aventis and TEVA.
R.G. has received personal compensation for activities with Bayer Healthcare, Biogen and Teva Neuroscience and in an editorial capacity from Therapeutic Advances in Neurological Disorders, and also received patent payments from Biogen and research support from Bayer Healthcare, Biogen, Merck Serono, Teva Neuroscience, Novartis and from the German Ministry for Education and Research (BMBF, “German Competence Network Multiple Sclerosis” (KKNMS), CONTROL MS, 01GI0914).
A.C. has received personal compensation for activities with Almirall Hermal GmbH, Bayer Schering, Biogen, Merck Serono, Novartis and Teva Neuroscience, research support from Bayer Schering, Biogen, Merck Serono and Novartis and research grants from the German Ministry for Education and Research (BMBF, “German Competence Network Multiple Sclerosis” (KKNMS), CONTROL MS, 01GI0914).
The other authors report no disclosures.
Competing interests
The authors declare that they have no competing interests.
Authors’ contributions
Study concept and design: Chan, Gold, Lill Performance of animal experiments (in vivo, ex vivo): Schrewe, Demir, Böhme Acquisition of human data: Lill, Gerdes, Guillot-Noel, Akkad, Blaschke, Graetz, Hoffjan, Kroner, Rieckmann, Cournu-Rebeix, Zipp, Kümpfel, Buttmann, Zettl, Fontaine, Bertram, Chan Analysis and interpretation of data: Schrewe, Lill, Liu, Bertram, Chan, Salmen, Demir Drafting of manuscript: Schrewe, Lill, Chan, Salmen Critical revision of the manuscript for important intellectual content: Gold, Hermann, Hagemann, ElAli, Gerdes, Guillot-Noel, Akkad, Blaschke, Graetz, Hoffjan, Kroner, Rieckmann, Cournu-Rebeix, Zipp, Kümpfel, Buttmann, Zettl, Fontaine, Bertram Administrative, technical and material support: Chan, Hermann, Hagemann, ElAli, Demir, Böhme All authors read and approved the final manuscript.