Citrullination of histone H3 interferes with HP1-mediated transcriptional repression.

Multiple Sclerosis (MS) is an autoimmune disease associated with abnormal expression of a subset of cytokines, resulting in inappropriate T-lymphocyte activation and uncontrolled immune response. A key issue in the field is the need to understand why these cytokines are transcriptionally activated in the patients. Here, we have examined several transcription units subject to pathological reactivation in MS, including the TNFα and IL8 cytokine genes and also several Human Endogenous RetroViruses (HERVs). We find that both the immune genes and the HERVs require the heterochromatin protein HP1α for their transcriptional repression. We further show that the Peptidylarginine Deiminase 4 (PADI4), an enzyme with a suspected role in MS, weakens the binding of HP1α to tri-methylated histone H3 lysine 9 by citrullinating histone H3 arginine 8. The resulting de-repression of both cytokines and HERVs can be reversed with the PADI-inhibitor Cl-amidine. Finally, we show that in peripheral blood mononuclear cells (PBMCs) from MS patients, the promoters of TNFα, and several HERVs share a deficit in HP1α recruitment and an augmented accumulation of histone H3 with a double citrulline 8 tri-methyl lysine 9 modifications. Thus, our study provides compelling evidence that HP1α and PADI4 are regulators of both immune genes and HERVs, and that multiple events of transcriptional reactivation in MS patients can be explained by the deficiency of a single mechanism of gene silencing.

Effects of interferon-beta therapy on elements in the antiviral immune response towards the human herpesviruses EBV, HSV, and VZV, and to the human endogenous retroviruses HERV-H and HERV-W in multiple sclerosis.

Effects of treatment of multiple sclerosis patients with IFN-ß on elements of the antiviral immune response to herpesviruses were analysed in a longitudinal study. We found significantly increased seroreactivity to EBV EBNA-1, and to VZV, in patients who did not respond to IFN-ß therapy. We found no significant changes in seroreactivity to EBV EA, or to HSV. For the same patient cohort, we have previously demonstrated significant decreases in seroreactivities to envelope antigens for the two human endogenous retroviruses HERV-H and HERV-W, closely linked to efficacy of therapy. We further searched for correlations between seroreactivities to EBV, HSV, and VZV, and levels of mannan-binding lectin (MBL), and MBL-associated serine protease 3. We found no such correlations. Our results are in accord with recent reports of increased seroreactivity to EBV EBNA-1, and to VZV in active MS, and they support that the herpesviruses EBV and VZV together with HERV-H/HERV-W and the antiviral immune response may play a role in MS development.

Expression of HERV-Fc1, a human endogenous retrovirus, is increased in patients with active multiple sclerosis.

Multiple sclerosis (MS) is considered to be an autoimmune disease with an unknown cause and with immune system dysregulation. Among environmental factors, viruses are most often connected with the etiology of MS. Human endogenous retroviruses (HERVs) constitute 5 to 8% of human genomic DNA and have been detected as transcripts and proteins in the central nervous system (CNS) and peripheral blood, frequently in the context of neuroinflammation. HERV-Fc1, which belongs to the HERV-H/F family, has received our attention largely because of the genetic association with MS. We studied the expression of a capsid (Gag) protein of HERV-H/F origin by flow cytometry in peripheral blood mononuclear cells (PBMCs) from healthy controls and from MS patients with nonactive or active disease. There was a significant increase in HERV-H/F Gag expression in CD4(+) (P < 0.001) and CD8(+) (P < 0.001) T lymphocytes and in monocytes (P = 0.0356) in PBMCs from MS patients with active disease. Furthermore, we have undertaken the first rigorous SYBR green-based absolute quantitative PCR (Q-PCR) evaluation approach to quantify extracellular HERV-Fc1 RNA viral loads in plasma from MS patients and healthy controls. We found a 4-fold increase in extracellular HERV-Fc1 RNA titers in patients with active MS compared with healthy controls (P < 0.001). These findings strengthen the link between HERV-Fc1 and the pathology of MS. The cause and biological consequences of these differential expression levels will be the subject of further investigation. HERV-Fc1 biology could be a compelling area for understanding the pathology of MS and possibly other autoimmune disorders.

The etiology of multiple sclerosis: genetic evidence for the involvement of the human endogenous retrovirus HERV-Fc1.

We have investigated the role of human endogenous retroviruses in multiple sclerosis by analyzing the DNA of patients and controls in 4 cohorts for associations between multiple sclerosis and polymorphisms near viral restriction genes or near endogenous retroviral loci with one or more intact or almost-intact genes. We found that SNPs in the gene TRIM5 were inversely correlated with disease. Conversely, SNPs around one retroviral locus, HERV-Fc1, showed a highly significant association with disease. The latter association was limited to a narrow region that contains no other known genes. We conclude that HERV-Fc1 and TRIM5 play a role in the etiology of multiple sclerosis. If these results are confirmed, they point to new modes of treatment for multiple sclerosis.

Short-term exposure to human cytomegalovirus-infected fibroblasts induces a proportional increase of active CD94/NKG2A(+) natural killer cells.

Natural killer (NK) cells are essential components of the immune response against human cytomegalovirus (HCMV). As NK cells are part of the innate immune system providing an immediate defense against pathogens, short-term exposure to HCMV-infected cells may induce changes in the phenotype and function of these cells. To identify immediate reactions of NK cells to HCMV, we co-cultured peripheral blood mononuclear cells with HCMV-infected fibroblasts for 24 and 72 hours. A distinct, HCMV-mediated, proportional enlargement of a subset of NK cells expressing CD94/NKG2A was sustained throughout the period of incubation. As preceding studies have shown that HCMV can cause an increase in CD94/NKG2C(+) NK cells, our results were surprising. The NK cells showed intense upregulation of the early activation marker CD69 in response to HCMV. The CD94/NKG2A(+) NK cells demonstrated the highest expression of CD69. Studies of HCMV-induced interferon-gamma expression after 24 hours of co-culture showed that this cytokine was almost exclusively produced by the CD94/NKG2A(+) subset of NK cells. In summary, our data demonstrate that HCMV induces an immediate proportional enlargement of a functionally active CD94/NKG2A expressing subset of NK cells.

B cells and monocytes from patients with active multiple sclerosis exhibit increased surface expression of both HERV-H Env and HERV-W Env, accompanied by increased seroreactivity.

BACKGROUND: The etiology of the neurogenerative disease multiple sclerosis (MS) is unknown. The leading hypotheses suggest that MS is the result of exposure of genetically susceptible individuals to certain environmental factor(s). Herpesviruses and human endogenous retroviruses (HERVs) represent potentially important factors in MS development. Herpesviruses can activate HERVs, and HERVs are activated in MS patients. RESULTS: Using flow cytometry, we have analyzed HERV-H Env and HERV-W Env epitope expression on the surface of PBMCs from MS patients with active and stable disease, and from control individuals. We have also analyzed serum antibody levels to the expressed HERV-H and HERV-W Env epitopes. We found a significantly higher expression of HERV-H and HERV-W Env epitopes on B cells and monocytes from patients with active MS compared with patients with stable MS or control individuals. Furthermore, patients with active disease had relatively higher numbers of B cells in the PBMC population, and higher antibody reactivities towards HERV-H Env and HERV-W Env epitopes. The higher antibody reactivities in sera from patients with active MS correlate with the higher levels of HERV-H Env and HERV-W Env expression on B cells and monocytes. We did not find such correlations for stable MS patients or for controls. CONCLUSION: These findings indicate that both HERV-H Env and HERV-W Env are expressed in higher quantities on the surface of B cells and monocytes in patients with active MS, and that the expression of these proteins may be associated with exacerbation of the disease.

Effects of interferon-beta therapy on innate and adaptive immune responses to the human endogenous retroviruses HERV-H and HERV-W, cytokine production, and the lectin complement activation pathway in multiple sclerosis.

The effects of treatment of multiple sclerosis patients with IFN-beta on elements in the innate and adaptive immune response were analysed in a longitudinal study. We demonstrate significant decreases in anti-Envelope antibody reactivity for the two closely related Gammaretroviral human endogenous retroviruses (HERVs), HERV-H and HERV-W, as a consequence of IFN-beta therapy, closely linked to efficacy of therapy/low disease activity. We also show strong indications of a protective effect of high levels of two components in the innate pathogen-associated molecular pattern recognition: mannan-binding lectin (MBL), and MBL-associated serine protease 3 (MASP-3). Serum levels of typical Th1- and Th2-related, MS-relevant cytokines were also monitored. Overall both Th1- and Th2-associated cytokines were modestly, albeit significantly up-regulated, notably IL-2 and TNF-alpha (MS patients with inactive disease), as well as IL-4 and, to some extent IL-10 (no increase in IL-10 for MS patients with active disease (non-responders)). We found no overall changes in Th1/Th2 ratios. Our results support that HERV-H/HERV-W and the antiviral immune response may play a role in MS development, and that these HERVs have potential as biomarkers for disease activity.

Activation of endogenous retrovirus reverse transcriptase in multiple sclerosis patient lymphocytes by inactivated HSV-1, HHV-6 and VZV.

Human endogenous retroviruses (HERVs) and herpesviruses have been associated with the development of multiple sclerosis (MS). These virus groups interact with each other and have been shown to induce synergistic immune responses. Here, we focus on the possible role of herpesviruses as contributing factors in HERV activation. We demonstrate the ability of HSV-1, HHV-6, and VZV antigens to induce higher RT activity in peripheral lymphocytes from MS patients vs. controls during the first 6 days post-antigen stimulation. On subsequent days, only VZV can sustain the increase in the RT expression in cells from MS patients. The RT induction does not depend on herpes replication.

Gene-environment interactions in multiple sclerosis: innate and adaptive immune responses to human endogenous retrovirus and herpesvirus antigens and the lectin complement activation pathway.

Aspects of gene-environment interactions in multiple sclerosis (MS) were analysed in serum samples from 46 MS families (25 sporadic MS cases and 42 familial MS cases): antibodies to the MS-associated human endogenous retrovirus HERV-H, and levels of three components in the innate pathogen-associated molecular pattern recognition: mannan-binding lectin (MBL), and MASP-2 and MASP-3. For representative MS families, we also determined herpesvirus serology for HSV-1, VZV, and EBV; and tissue typed for HLA-B, and HLA DR and DQ. In MS, a significant correlation between elevated immune reactivity to HERV-H Env and disease activity was demonstrated, as were indications of a protective effect of high MBL and MASP-3 levels. The HLA alleles B*07, DRB*02, and DQB1*06 were commonly present together in the MS families, both in MS patients, and in unaffected family members. Our results support that HERV-H and the antiviral immune response may play a role in MS development, and also underline the tenuous nature of specific genetic contributions to this complex disease.

Simultaneous presence of endogenous retrovirus and herpes virus antigens has profound effect on cell-mediated immune responses: implications for multiple sclerosis.

Retroviruses have been suggested as possible pathogenic factors in multiple sclerosis (MS), supported by the observation that endogenous retroviruses are activated in MS patients. Different members of the herpes family of which several are neurotropic have also been suggested as factors in MS pathogenesis. Further, interactions between retroviruses and herpes viruses have been implied in the development of MS. The objective of the study was investigation of cell-mediated immune responses of MS patients to retrovirus and herpes virus antigens, particularly antigen combinations, with analyses of the influence of retrovirus antigens on cellular immunological reactivity toward other viral antigens. Cellular immunity as measured by blast transformation assays was analyzed using freshly isolated peripheral blood mononuclear cells from 47 MS patients and 36 healthy volunteers. Combinations of the endogenous retrovirus HERV-H and herpes virus antigens resulted in highly increased cellular immune responses among both the MS patients and healthy subjects. The increase was synergistic in character in most samples. Very pronounced effects were obtained using HHV-6A and HSV-1 antigens. Blast transformation assays combining antigens from two different herpes viruses or combinations of measles and herpes antigens showed no synergy. The obtained data indicate a pronounced synergistic effect on the cellular immune response when retrovirus and herpes antigens are present together. The cause of the synergy is unknown so far. The effect on the immune response may influence the disease progression.

[Human endogenous retroviruses and disease?]. / Humane endogene retrovirus og sygdom?

Endogenous retroviruses represent sequences descended from ancient virus infections integrated in the host genome. They participate in processes, such as speciation, recombination, ontogenesis, and regulation of tissue specificity and gene expression. It has been suggested in recent years that human endogenous retroviruses may play a role in certain types of cancer and autoimmune diseases. Human endogenous retroviruses represent both putative susceptibility genes and putative pathogenic viruses in diseases like systemic lupus erythematosus, Sjögren's disease, rheumatoid arthritis, and possibly type 1 diabetes. Multiple sclerosis is specifically associated with expression of human endogenous retroviruses as virions. It is not yet known if the human endogenous retroviruses also represent causal factors, but several pathogenic mechanisms are possible.

A transmissible human endogenous retrovirus.

The transmissibility of the human endogenous retrovirus HERV-H/RGH-2 was investigated by marker rescue: intraspecies transmission of HERV-H/RGH-2 retrovirus particles was attempted by cocultivation of virion-producing, long-term cell cultures spontaneously formed from peripheral blood mononuclear cells from several multiple sclerosis patient cultures with a retroviral vector construct-harboring cell line. Transmissibility was assessed by assays for productive infection (reverse transcriptase activity), and assays for rescue of the retroviral vector construct in indicator cells. Our studies show that the human endogenous retrovirus HERV-H/RGH-2 is transmissible, albeit at a very low level. The human endogenous retrovirus HERV-H/RGH-2 is associated with multiple sclerosis (MS). Previously, we demonstrated sequence variants of the human endogenous retrovirus HERV-H family in virion form, by applying RT-PCR to virion RNA from the supernatants of long-term MS cell cultures, and to the particulate fraction of a series of MS patient plasma samples.