Op het forum van Phoenix Rising is de posterpresentatie

van prof. de Meirleir en kollega's tijdens de eerste internationale XMRV-workshop gepubliceerd.

Alhoewel de studie zeer klein was (16 XMRV-kweekkultuur-positieve ME/CVS-patiënten),

bevestigen de resultaten van deze studies alle kenmerkende biologische afwijkingen

(inflammatie, immuundysfunctie/-suppressie, lekke darm en translokatie van darmbakteria).

Abnormaliteiten die al jaren door "vermoeidheidsdeskundigen" ontkend of genegeerd worden.

De voornaamste bevindingen van de Meirleir, Metzger, Roelant en Frémont in een notedop:

• Het aantal witte bloedcellen met de CD3+ (rijpe T-cellen) en CD57+ marker zijn verlaagd,
• de koncentraties tussenproduct c4a van het komplementsysteem (dat gericht is op het opruimen van met antistoffen gemarkeerde cellen en in een studie van Nijs et al. gezien wordt als een marker voor post-exertional malaise) en elastase zijn beduidend hoger,
• de koncentratie met CD14 gemarkeerde witte bloed-cellen in het serum (dat gezien wordt als een maatstaf voor de aanwezigheid van lipopolysacchariden, endotoxinen van gram-negatieve [darm]bakteria, die een sterke immuunresponse veroorzaken) is veel hoger,
• de hoeveelheid immunoglobuline IgG3 in het serum is verlaagd: IgG3 subklasse-deficiëntie,
• de cytokines IL-8 en IL-10 en chemokines MCP-1 en MIP-1b zijn (sterk) toegenomen en vormen een mogelijke immunologische vingerafdruk van een XMRV-infektie

(zoals eerder vastgesteld in een studie van Lombardi, Mikovits en kollega's),

• de koncentraties IgA (in hoge mate verantwoordelijk voor de eerste lijn van afweer in de darmen) in de ontlasting is verlaagd (indikatief voor schade aan het slijmvlies?).

Voor de PDF-versie van de posterpresentatie, klik op onderstaande afbeelding:

Is the mechanism of systemic immune activation in XMRV positive CFS patients similar to that observed in HIV?

Kenny De Meirleir M.D., PhD, Vrije Universiteit Brussel, Brussel, Belgium;

Marc Frémont, PhD;

K. Metzger, MS;

C. Roelant, PhD, RED Laboratories NV, Zellik, Belgium

BACKGROUND

Chronic activation of the immune system

is a hallmark of progressive HIV infection and

is a better predictor of disease outcome than plasma viral load.

Brenchley et al showed that

circulating microbial products

were a cause of HIV-related immune activation,

which they called microbial translocation.

We have previously shown that

Chronic Fatigue Syndrome (CFS) patients suffer

from gastrointestinal dysbiosis 2 and

from immune dysfunctions 3.

Because of the recent discovery of

presence of an infectious retrovirus, XMRV

in blood cells of patients with chronic fatigue syndrome

by Lombardi et al 4,

we wanted to test the hypothesis that

the pathophysiology of the systemic immune activation

in XMRV positive CFS patients

could be similar to the one observed in HIV.

PATIENTS AND METHODS

Sixteen CFS patients fulfilling the Canadian criteria 5

who were found positive for XMRV by co-culture technique 4

(a sensitive cell culture assay for detection of XMRV)

were included in the study.

Because of the low number of subjects studied

we chose to use reference data from a large control population

which were collected earlier.

Immunophenotyping was performed in the laboratory of clinical hematology

(University Hospital Brussel, Belgium)

using standard flow cytometry:

• T cells (CD3+, CD4+, CD8+, CD4/CD8, CD3+CD16CD56+),
• B cells (CD19+),
• NK cells (CD3-CD16CD56+),
• other (CD2+, CD2+CD25+, CD25+, CD3+HLADR+, CD19+CD5+, CD57+).

Elastase activity was measured in monocytes and lymphocytes

using an enzymatic colorimetric assay EnzChek® Elastase Assay kit E-12056

(Molecular Probes, OR, USA).

C4a: an elisa technique was used using Becton Dickinson OptElA human C4a elisa kit.

lgG3: nephelometry in serum.

Cytokines and sCD14: serum level measurement

using Becton Dickinson Cytometric Bead Assay system.

Perform: mRNA level measured by real-time PCR,

using gene expression assay reagents from Applied Biosystems.

Stool IgA: sent to Diagnos-Techs (Seattle, Washington, and analysed in their labs).

The descriptive statistics for the different parameters are presented in a table on this poster.

STATISTICAL ANALYSIS

A one sided t-test was used

to test the hypothesis that

the mean value of the XMRV group

is significantly different from the middle of the normal range

(representing normal population).

A two-sided test was used

when abnormal values can occur at both sides of the normal range,

otherwise a one-sided test was used.

The significance level was set at 0.05.

The statistical analysis was performed by Prof. D. Coomans

at the department of medical statistics of

the faculty of medicine and pharmacy at the Vrije Universiteit Brussel.

RESULTS

The number of CD3+ T cells and CD57+ lymphocytes

was significantly lower

compared to the reference values.

C4a and elastase activity

were significantly higher

in the XMRV positive CFS population.

Soluble CD14

which codes for LPS in the plasma

was significantly higher

at p < 0.001 compared to the reference population.

XMRV positive CFS patients

had significantly higher

serum IL-10, MCP-1 , MIP-1 beta and IL-8 levels.

Serum levels of other cytokines

(IL-12, IL-1 beta, IL-6, TGF-beta and alpha-TNF)

were not statistically different

compared to the reference values.

Other lymphocyte subsets

showed

no difference from the reference in the XMRV positive patients.

Stool lgA and lgG3

were statistically lower

in the XMRV positive patients.

CONCLUSION

The results of this preliminary study in a limited number of subjects

show that XMRV positive CFS patients have

lower than normal levels of lymphocytes and low numbers of CD57+ lymphocyte subtype

as in HIV.

The absolute numbers of CD4+ and CD8+T cells

were not statistically different from the reference values,

but expanding this study to a larger number of patients is necessary

to make solid statements in this regard.

XMRV-positive CFS patients have an activated innate immune system

(elastase activity, increased C4a)

which could be related to microbial translocation

as their sCD14 is significantly higher than expected;

sCD14 strongly correlates with plasma LPS 1.

Low stool IgA (in some of these 16 patients it was undetectable)

also points towards a dysfunctional mucosa-associated lymphomal tissue (MALT)

in XMRV-positive CFS patients.

Furthermore we found that

these patients as a group have lower than normal lgG3 serum levels.

The cytokines IL-8, IL-10, MCP-1 and MIP-1beta are increased

and might constitute a biological signature for the viral infection.

These observations and others

(unpublished data on serum levels of LPS in CFS patients)

provide evidence for

microbial translocation

being part of

the pathophysiology

of XMRV positive patients.

REFERENCES

1. Brenchley J.M., et al. Microbial translocation is a cause of systemic immune activation in chronic HIV infection. Nature Medicine 12 (12): 1365-1371, 2006.
2. Fremont M., et al. Detection of herpes-viruses and parvovirus B19 in gastric and intestinal mucosa of chronic fatigue syndrome patients. In Vivo, 23: 209-214, 2009.
3. Meeus M., et al. Unraveling Intracellular Immune Dysfunctions in Chronic Fatigue Syndrome: Interactions between Protein Kinase R Activity, RNase-L cleavage and Elastase Activity and their Clinical Relevance. In Vivo 22: 115-122, 2008.
4. Lombardi V., et al. Detection of an infectious retrovirus, XMRV, in blood cells of patients with Chronic Fatigue Syndrome. Science 326: 585-589, 2009.
5. Carruthers B, et al. Myalgic encephalomyelitis / chronic fatigue syndrome: Clinical working case definition, diagnostic and treatment protocols. J. Chronic Fatigue Syndrome 11: 1-12, 2003.

Met dank aan Terry die mij opmerkzaam maakte op het dokument.