Assessing cellular energy dysfunction in CFS/ME using a commercially available laboratory test, by Cara Tomas, Tiffany A Lodge, Michelle Potter, Joanna L Elson, Julia L Newton, Karl J Morten, in Sci Rep. 2019 Aug 7;9(1):11464. doi: 10.1038/s41598-019-47966-z.
Research abstract:
The mitochondrial energy score (MES) protocol, developed by the Myhill group, is marketed as a diagnostic test for chronic fatigue syndrome/Myalgic Encephalomyelitis (CFS/ME). This study assessed the reliability and reproducibility of the test, currently provided by private clinics, to assess its potential to be developed as an NHS accredited laboratory test. We replicated the MES protocol using neutrophils and peripheral blood mononuclear cells (PBMCs) from CFS/ME patients (10) and healthy controls (13). The protocol was then repeated in PBMCs and neutrophils from healthy controls to investigate the effect of delayed sample processing time used by the Myhill group.
Experiments using the established protocol showed no differences between CFS/ME patients and healthy controls in any of the components of the MES (p ≥ 0.059). Delaying blood sample processing by 24 hours (well within the 72 hour time frame quoted by the Myhill group) significantly altered many of the parameters used to calculate the MES in both neutrophils and PBMCs. The MES test does not have the reliability and reproducibility required of a diagnostic test and therefore should not currently be offered as a diagnostic test for CFS/ME. The differences observed by the Myhill group may be down to differences in sample processing time between cohorts.
Discussion
Given the evidence presented here, we advise that the MES test should not be used as a diagnostic test in its current form as in this study shows there to be no differences between CFS/ME and control results when the MES protocol was followed using fresh blood samples. This is contrary to results from the group who devised the test and offer it to patients.
We explored the impact of delayed sample processing on blood glucose concentration in the collection tube as a possible explanation for the discrepancies in results between our group and the Myhill group. As expected with such high cell numbers the glucose rapidly dropped as the cells utilized the glucose. In addition the neutrophil component on FACS analysis in the white cell fraction showed differences in size and granularity between the 1 hr and 24 hr fractions suggestive of altered properties. Having taken into account a 24 hour delay between blood collection and cell isolation, we have shown decreases in ATP parameters in control cells similar to those seen by the Myhill group in the CFS/ME patients. We suggest that it is potentially the delay between sample collection and cell isolation that is causing the decrease in mitochondrial function previously reported in CFS/ME patients.
While this study used relatively small samples sizes compared with the original study, abnormalities in CFS/ME patients should be reproducible even in small sample sizes given the current use of this test for diagnostic purposes.
The Myhill group have recently altered their protocol to use PBMCs instead of neutrophils, however, this research has not been published and we have no information on the control ranges used and whether they were developed from blood samples processed over 24 hrs. The CFS/ME PBMC study by Tomas et al. did show the utility of using PBMC using the Seahorse extracellular flux analyser to study energetics3. However, the sudden switch to using PBMCs for the MES protocol appears to be as a result of criticism over the use of neutrophils rather than being an evidence lead change. There has been no data published from the Myhill group regarding the suitability of PBMCs with their previously established protocol, or any publication of results with the new cell type. After a diagnosis of CFS/ME is made using the MES test, patients are subsequently sold supplements in order to treat their CFS/ME, despite there being no placebo-controlled trial to show their effectiveness. The first peer-reviewed publication regarding the MES test from the Myhill group came after they had already been using the test and supplement regime with CFS/ME patients despite there being no published evidenced of the effectiveness, reliability, or reproducibility of the test. Additionally, the MES test has not been conducted using other patient groups with fatigue as a core symptom, therefore its specificity for CFS/ME has not been confirmed.
If energetic dysfunction is to be used as a marker of CFS/ME its exact role in the condition needs to be better understood including studying energetic dysfunction in other fatigue groups. Only when we have clearer understanding of the disease process and knowledge of specific factors shown to be different in CFS/ME should we consider developing a diagnostic test to aid in treatment strategies and determining outcome in clinical trials.
Clinicians approached by patients with results from the MES test should be advised to interpret the results with caution, while patients considering paying for the test should be advised of the lack of supporting scientific evidence. The test in its current form does not have the reliability or reproducibility required of a diagnostic test and therefore should not be offered by the NHS or private clinics as a diagnostic test for CFS/ME. Other tests of energetic dysfunction could be developed using the seahorse extracellular flux assay but more research is required as to the meaning of the results in the aetiology of CFS/ME before a test using this approach should be developed.
Comment from the funders of the study, the ME Association: Independent researchers determine mitochondrial test is unreliable and should not be used as test in ME/CFS
Responding to criticism of their mitochondrial test Myhill & McLaren emphasize that the test has “never been presented as a diagnostic test for CFS/ME.” They query whether the researchers did a true replication as they had offered to do a comparison test in their labs on the same samples but the offer wasn’t accepted. Regarding the technical details, Dr McLaren says he finds it strange that no significant differences were found between fresh & frozen cells, which is at odds with what he & other scientists have found while exploring the test.
