Tuesday, 6 February 2018

That damned, elusive EBV

Will this study get the field behind the EBV hypothesis once and for all? Or will it be another bun fight?




PLoS One. 2018 Feb 2;13(2):e0192109. doi: 10.1371/journal.pone.0192109. eCollection 2018.

Epstein-Barr virus is present in the brain of most cases of multiple sclerosis and may engage more than just B cells.

Hassani A, Corboy JR, Al-Salam S, Khan G.

Abstract


Multiple sclerosis (MS) is a chronic neuroinflammatory condition of the central nervous system (CNS). It is a major cause of neurological disability in young adults, particularly women. What triggers the destruction of myelin sheaths covering nerve fibres is unknown. Both genetic and infectious agents have been implicated. Of the infectious agents, Epstein-Barr virus (EBV), a common herpesvirus, has the strongest epidemiological and serological evidence. However, the presence of EBV in the CNS and demonstration of the underlying mechanism(s) linking EBV to the pathogenesis of MS remain to be elucidated. We aimed at understanding the contribution of EBV infection in the pathology of MS. We examined 1055 specimens (440 DNA samples and 615 brain tissues) from 101 MS and 21 non-MS cases for the presence of EBV using PCR and EBER-in situ hybridization (EBER-ISH). EBV was detected by PCR and/or EBER-ISH in 91/101 (90%) of MS cases compared to only 5/21 (24%) of non-MS cases with other neuropathologies. None of the samples were PCR positive for other common herpesviruses (HSV-1, CMV, HHV-6). By quantitative PCR, EBV viral load in MS brain was mainly low to moderate in most cases. However, in 18/101 (18%) of MS cases, widespread but scattered presence of EBV infected cells was noted in the affected tissues by EBER-ISH. Immunohistochemical analysis of EBV gene expression in the 18 heavily infected cases, revealed that the EBV latent protein EBNA1, and to a lesser extent the early lytic protein BZLF1 were expressed. Furthermore, using double-staining we show for the first time that astrocytes and microglia, in addition to B-cells can also be infected. To the best of our knowledge, this is the most comprehensive study demonstrating that EBV is present and transcriptionally active in the brain of most cases of MS and supports a role for the virus in MS pathogenesis. Further studies are required to address the mechanism of EBV involvement in MS pathology.



"We seek him here, we seek him there...Is he in heaven - Is he in hell?"
-The Scarlett Pimpernel by E Orczy


No really! The cause of MS is as elusive as the Pimpernel was to the French. Despite mounting evidence, no research group has yet successfully managed to nail EBV to MS. This work by Hassani and colleagues is hope that it may yet happen!

Epstein-Barr virus (EBV) is a member of the Herpes family of viruses and is common in humans. It's target is B cells and can remain dormant or latent in memory B cells for their lifespan. A small percentage, however, do reactivate in what is called the lytic phase.

EBV is associated with more than one disorder, but is thought in MS at least it causes a virus-host immune system response imbalance, with the immune response to the virus to the virus itself being disrupted (Sundström P, Juto P, Wadell G, Hallmans G, Svenningsson A, Nyström L, et al. An altered immune response to Epstein-Barr virus in multiple sclerosis: a prospective study. Neurology. 2004;62: 2277–2282). Almost all PwMS demonstrate evidence of previous EBV infection, and those who have not been exposed (i.e. seronegative) have almost zero risk of developing MS. The spanner in the works is that ~95% of those without MS have also had previous EBV exposure (Ascherio A, Munch M. Epstein-Barr virus and multiple sclerosis. Epidemiology. 2000;11: 220–224).


The question then becomes can you place EBV successfully in the brain where it can cause MS? And that is what the authors have done. Hassani et al, examined the presence of EBV in 122 post-mortem MS brains and non-MS brains. Others have tried to do this in the past but not successfully for any number of reasons - techniques, sampling etc. They used a PCR technique and a highly sensitive and specific EBER- in situ hydridization (EBER-ISH) technique to localise EBV infected cells in brain tissues.

Surprisingly, they found that EBV was present in 90% of MS cases, but also found no presence of other viruses, such as HSV-1, CMV and HHV-6 that have been previously implicated in MS. Of note, they also found that not only did EBV target B cells, but also infected ~10-15% activated microglia and astrocytes (the innate immune system, see Figure). Therefore, the role of EBV in the inflammatory cascade needs to be looked at in greater detail.

Overall, a very interesting paper and others will now have to re-examine EBV in MS brains using the more sensitive techniques in order to identify scattered and low level EBV infection.

Figure: Double staining for EBV and cellular markers.
Double staining for EBV (EBER-in situ hybridization: dark blue staining) and different cellular markers (immunohistochemistry: brown) in the white matter of 3 different heavily infected MS cases. The pattern of double-staining seen in these 3 cases is representative of that seen in other double-positive cases. (A) EBV and CD20 (B-cell marker), (B) EBV and GFAP (astrocyte marker), (C) EBV and Iba1 (microglia marker). Double positive cells are indicated by the arrows.

21 comments:

  1. "However, in 18/101 (18%) of MS cases, widespread but scattered presence of EBV infected cells was noted in the affected tissues by EBER-ISH"

    Does the paper pass the "smack you in the eyes" test?


    Obrigado

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    1. Many other gorups have looked and have not found the same high level but this group looked at multiple samples from the same person. Also the sensitivity was not one virus copy per cell and so the umber of infected could be higher and would need to be is it were causal

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  2. Is this study saying that the majority of MSers are CMV-free?

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    1. I doubt it. Most of us have CMV but in this study they don't find it in the brain

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  3. My doctor says that she has many EBV seronegative patients and I have met such cases myself. Even if there is a 5% of EBV seronegative MS patients, shouldnt that lead us to wider theories that would include EBV?

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    1. Many apparently EBV seronegatives on closer inspection are actually EBV positivewith more sensitive detection methods.

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    2. It is very important to confirm that.

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    3. I have MS and I'm EBV seronegative, blood taken at my hospital, requested by a neurologist.
      How do I tell if the detection method was not sensitive enough? Is the method on the results print out?

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    4. Serological detection will not be sensitive enough

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    5. It would be a relatively cheap study to see if the EBV- patients are truly EBV- and it could add a lot to the discussion.

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    6. Following on from this, in a study from team G "Only 1 of 1,047 patients (<0.01%) was truly EBV-negative."
      Paper can be viewed here;
      https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5292929/

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    7. How do pwMS request the more sensitive EBV blood test?
      Is it something that needs to be done privately rather than NHS? as it's not the standard EBV blood test.

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    8. In the case above it was not simply a more sensitive test it was just that they looked for immune responses against more EBV antigens, I dealy you would like to able to detect 1 copy of virus in a cell

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  4. What about the spinal cord? Is EBV also there? What is the correlation between EBV sites and lesions?

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    1. "What is the correlation between EBV sites and lesions?"

      It's been done.

      Molecular signature of Epstein-Barr virus infection in multiple sclerosis brain lesions

      "..although the presence of EBV-related proteins were detected in both MS and control brain samples, the significant prevalence of latent and lytic viral-associated proteins in MS brain samples, and especially in chronic inactive lesions suggest a key role of EBV during disease pathogenesis."

      http://onlinelibrary.ectrims-congress.eu/ectrims/2017/ACTRIMS-ECTRIMS2017/200600/may.h.han.molecular.signature.of.epstein-barr.virus.infection.in.multiple.html

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    2. Chronic inactive lesions are only a subset of total MS lesions. What is more, a putative causative role is strengthened by EBV findings in new and expanding lesions, not in old or inactive. Moreover, if EBV is abundant in non lesional territories also, then EBV traces inside lesions are far less meaningful. That is why a CNS-wide mapping of EBV sites is necessary.

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    3. Chronic inactive was once active..but thanks for your thoughts VV...student project anyone?

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  5. Don’t see why not, EBV can infect spinal cord and nerves. This study was focused on brains. EBV has been localised to meningeal lymphoid follicles and perivenular spaces, so there is a potential for correlation with lesions.

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  6. "why not", "potential".
    From these, i assume two things:
    1. Occurrence of EBV in MS spinal cord is still unknown.
    2. No mapping of EBV presence in brain and spinal cord exists yet. Consequently, no correlation with MS brain or spinal lesions yet.

    Therefore, a long way from calling it a cause, let alone drawing an immune mediated pattern of causation.

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  7. I just know that either I'm very "unlucky" or it was a "total coincidence" that I discovered MS soon after an IM relapse.

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