Khan F, Kennedy G, Spence VA, Newton DJ, Belch JJF
Vascular Diseases Research Unit, The Institute of Cardiovascular Research, Ninewells Hospital and Medical School, Dundee, UK
In the present study, we have investigated whether the peripheral cholinergic abnormalities that we have reported previously (Spence et al, Am J Med 2000; 108: 736–9) in patients with chronic fatigue syndrome (CFS) are also present in those with Gulf War syndrome (GWS) and agricultural workers exposed to organophosphate pesticides, where cholinesterase inhibition is specifically implicated. We also looked at whether these abnormalities might be due to a reduction in the activity of cholinesterase expressed on the vascular endothelium.
Methods and results
We used laser Doppler imaging to measure the forearm skin blood flow responses to iontophoresis of acetylcholine and of methacholine (which is resistant to breakdown by cholinesterase) in patients with CFS, GWS and those with a history of ill health after definite organophosphate exposure, as well as in matched healthy controls. The response to acetylcholine was significantly higher in patients with CFS than in controls (p=0.029, repeated-measures ANOVA), but was normal in those with GWS and those exposed to organophosphates. The methacholine response was higher than the acetylcholine response in all patient groups except for those with CFS, where there was no difference between the responses.
Although there are many clinical similarities between these three illnesses, our results indicate peripheral cholinergic abnormalities in the vascular endothelium of only patients with CFS, suggesting that this syndrome has a different aetiology, which might involve inhibition of vascular cholinesterase.
Acetylcholine abnormalities are particularly implicated in people such as Gulf War veterans and agricultural workers, who have been exposed to substances which inhibit the activity of the enzyme cholinesterase. These people suffer from symptoms which can be very similar to those of ME/CFS, although there are important differences.
We compared the acetylcholine blood flow responses of a group of these people with the responses of ME/CFS patients, to see whether they have the same abnormalities. However, the Gulf War veterans and agricultural workers all tended to have a normal response to acetylcholine, suggesting that their illnesses are fundamentally different to ME/CFS. This is interesting considering that patients from all of these groups are often collected together under the term ‘chronic fatigue syndrome’.
Methacholine is a chemical which is almost identical, and has similar effects, to acetylcholine, except that it is much less sensitive to the action of cholinesterase. For this reason, the blood flow response to methacholine is normally higher than that to acetylcholine. However, in patients with ME/CFS, they were similar; further evidence of a cholinesterase abnormality.
Comment by ME Research UK
This is the fourth paper in a series of studies looking at endothelial-dependent vasodilator mechanisms in patients with chronic fatigue syndrome (CFS) and related disorders. Other papers will follow in an attempt to unravel why the endothelium is sensitive to acetylcholine (ACh) in patients within the CFS construct, many of whom fit the description for myalgic encephalomyelitis (ME) and post-viral fatigue syndrome.
The first paper in the series (1) demonstrated that the microcirculation in CFS patients was sensitive to ACh but not to the endothelial-independent vasodilator sodium nitroprusside — a donor of nitric oxide (NO). (Read more here.) This sensitivity was unusual because all of the other diseases so far studied in this way show a blunted response to acetylcholine.
The second paper in the series (2) was on patients with a diagnosis of primary fibromyalgia, and in this study there was no evidence of sensitivity to either ACh or NO.
In subsequent experiments (3), we tried to determine why CFS/ME patients had this increased blood vessel sensitivity to ACh. We hypothesised that the sensitivity might be a consequence of a reduced expression of acetylcholinesterase (AChE) on the vascular endothelium — AChE is the enzyme which breaks down ACh. We tested this by monitoring the blood vessel response to ACh throughout its complete time-course; i.e., from baseline to peak flow and back to baseline. We found that the time taken for blood flow to return to normal following ACh stimulation was significantly longer than normal in CFS/ME patients, and this provided additional support for the hypothesis that the breakdown of the ACh response was somehow impaired.
In the paper just published, we now show two things: ACh-sensitivity is not a feature of patients with organophosphate-induced delayed neuropathy, nor is it a feature of veterans of the 1991 Gulf War conflict who have developed Gulf War Syndrome (GWS). These findings surprised us because we had hypothesised that the symptoms of both of these patient groups might be mediated by exposure to agents that inhibit AChE. We chose farmers who had a definite history of exposure to OPs, and we chose veterans with GWS who had taken the cholinesterase inhibiting agent pyridostigmine bromine (used prophylactically as nerve agent protection tablets) and who had also been in contact with OP insecticides used in the Gulf War theatre to delouse Iraqi captives. Despite the fact that we chose OP and GWS patients who fitted the CDC-1994 criteria for CFS, we found no evidence of sensitivity to ACh.
The second part of this new study looked at the endothelial response to methacholine (MCh).MCh is an endothelium-dependent vasodilator almost identical to ACh, with the exception that is much less affected by the catabolic actions of AChE. We predicted, therefore, that if the increased responsiveness to ACh in CFS/ME patients was a consequence of a reduced expression of vascular AChE then we would find no difference between the responses to ACh and MCh. This is indeed what we found. In control subjects, MCh responses were significantly greater than those for ACh, and this was also true for the OP and GWS cohorts.
These latest results are important for two reasons. Firstly, despite the fact that the symptoms of CFS/ME overlap considerably with those of OP-exposed and GWS patients, this is the first evidence to demonstrate that these syndromes are aetiologically different. Recent research has sought to include OP and GWS patients within the CFS construct, but this work published in Clinical Science serves as a reminder that grouping patients together on the basis of vague and non-specific symptoms will hinder research and make it much more difficult to unravel the cause of any of them. Secondly, this new work strengthens the hypothesis that AChE expressed on the endothelium is impaired in CFS/ME patients. It does not prove that under-expression of AChE is the cause of the ACh sensitivity — further experiments are now underway to test this — but it remains the best possible hypothesis on the basis of the experiments completed so far.
Finally, tests of ACh sensitivity are not diagnostic markers for CFS/ME. The vascular endothelium is a dynamic organism that is influenced by many biological factors, such as age, hormonal status, blood pressure, hyperlipidaemia and cardiovascular disease progression. So, it is at best a blunt instrument for the assessment of specific disturbances to cholinergic mechanisms. The findings reported here are specific to endothelial cholinergic activity and may or may not be applicable to other more widespread neurotransmitter functions of ACh. Nevertheless, the results reported in Clinical Science are unusual, if not unique to CFS/ME, and such sensitivity may well be important in terms of abnormal vascular control mechanisms that seem so characteristic of most CFS/ME patients.
- Spence VA, Khan K, Belch JJF. Enhanced sensitivity of the peripheral cholinergic vascular response in patients with chronic fatigue syndrome. Am J Med 2000; 108: 736–9.
- Al-Allaf AW, Khan F, Moreland J, Belch JJF, Pullar T. Investigation of cutaneous microvascular activity and flare response in patients with fibromyalgia syndrome. Rheumatology 2001; 40: 1097–1101.
- Khan F, Spence VA, Kennedy G, Belch JJF. Prolonged acetylcholine-induced vasodilatation in the peripheral microcirculation of patients with chronic fatigue syndrome. Clinical Physiology and Functional Imaging 2003; 23: 282–5.
Read more (pdf 155 KB) in the Spring 2005 issue of Breakthrough.