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CFS and FMS Research
Abstracts presented at the Sydney ‘98 and ‘99 International CFS Conferences
[Aphthous ulceration and symptoms in polysymptomatic patients]
[Alterations in plasma lipid composition in patients with CFS]
[The development of laboratory tests for investigating CFS and pain disorders - 1998]
[Chronic facial muscle pain and dysregulated cellular proteolysis -1998]
[Assessment of lipid homeostasis in sudden and gradual onset CFS patients - 1998]
[Alterations in urinary amino and organic acid excretion in patients with CFS - 1998]
[Chlorinated pesticides and chronic fatigue related illness]
[Chronic pain and protein turnover in polysymptomatic patients]
HOST VERSUS ACQUIRED RESPONSES IN DEFINED CFS PATIENTS
McGregor NR, Dunstan RH, Butt HL, Roberts TK, Klineberg IJ
The current CFS definitions constitute a heterogeneous (or diverse) group of conditions. CFS symptoms can be divided into basic two categories:
1. Those related to the host response (fatigue, pain, cognitive alterations).
2. Symptoms that indicate an acquired or infectious response (fever, sore throat, lymphodynia). © Bioscreen, CPRU, S Ashton and F Bartosy 1999 - 2001
Common host responses such as fatigue and pain are of multiple types and can be initiated by many different mechanisms. The use of these highly variable responses, such as fatigue or pain, as diagnostic markers of any disease is philosophically flawed as they give little indication of the potential aetiology(s).
Therefore analysis of those features associated with the acquired or infectious responses may allow identification of the potential aetiology of a number of conditions currently defined as CFS.
The analysis of the host response may allow determination of genetic or acquired susceptibility to the various acquired conditions.
Analysis of plasma lipids and urinary organic and amino acid excretion, along with symptom presentation allow the detection of both acquired and host responses. These could be used to differentiate between the various potential disease entities.
The patient’s disease status may be influenced by a combination of the following factors:
1. Auto-immune and persistent viral conditions.
2. Chronic bacterial infections/toxicity and environmental toxin exposure.
2. Secondary viral reactivation along with host genetic and acquired susceptibility.
This information clearly indicates the heterogeneous nature of the disease entities currently defined by the CFS definition. The use of CFS as a clinical diagnosis is not supported by the available data and should be replaced with the diagnosis of a chronic fatigue disorder.
APHTHOUS ULCERATION AND SYMPTOMS IN POLYSYMPTOMATIC PATIENTS
N R McGregor, H L Butt, R H Dunstan, T K Roberts, I J Klineberg
The reporting of a sore throat in patients complying with the CFS definition is often associated with oral and throat recurrent aphthous ulceration (RAU). Current evidence suggests that RAU represents reactivation of viruses such as CMV, Varicella zoster, enterovirus or HSV 6, but not HSV 1.
To evaluate the incidence of RAU in chronic fatigue/pain patients, 115 non-CFS and 204 CFS polysymptomatic patients were assessed for aphthous ulceration. It was found that 26.9% of the polysymptomatic patients had recurrent oral ulceration consistent with RAU, compared to 24.6% of control subjects.
This data indicated that the incidence of RAU is the about the same in the control subjects as in the polysymptomatic patients. A more extensive evaluation was made with the polysymptomatic patient group in comparison to a smaller control group (n=32) which had a more extensive array of symptom evaluations and pathology investigations. At the time of examination, active RAU was noted in 5.9% of control subjects compared to 14.7% of non-CFS polysymptomatic patients and 33.8% of CFS patients.
RAU was associated with very similar symptoms in both CFS and non-CFS patients. There were increases in the prevalence of history of spinal disk prolapse problems and appendectomy, and increases in the prevalence and/or severity of low back pain, cervical Iymphodynia, palpitations, muscle cramps, problems with concentration, loss of libido, as well as reduced gastric motility. Reductions in urinary frequency and the symptoms of irritable bowel were noted with alterations in both ulcer prevalence and severity.
Two potential disease models exist:
1. That CFS patients with RAU have a severe form of an RAU associated disease, such as Behcet's Syndrome.
2. That viral reactivation is a secondary phenomenon.
The RAU/symptom associations suggest that whilst CFS patients had an equal prevalence of history of RAU, they had an increased prevalence of active RAU. This strongly suggests that increased reactivation of existing viral infections occurs in CFS patients. Therefore, the symptoms associated with these reactivated viruses not only increase the patient's symptom prevalence and severity, but may also tend to confound the potential diagnosis of the underlying aetiology.
ALTERATIONS IN PLASMA LIPID COMPOSITION IN PATIENTS WITH CFS
Neil McGregor, Hugh Dunstan, Jennifer Watkins, Mark Donohoe, Timothy Roberts, Henry Butt, Raymond Murdoch, Warren Taylor
A study of 60 CDC-defined CFS patients and 39 age/sex matched controls was undertaken to determine whether the plasma lipid composition of CFS patients was different to that measured in control subjects.
It was found that there were no differences in the total concentrations of plasma lipids between the two groups. Multivariant analyses revealed that the total plasma concentrations were associated with cholesterol synthesis and lipogenesis in both CFS and control subjects. © Bioscreen, CPRU, S Ashton and F Bartosy 1999 - 2001
Alterations in omega 6 essential fatty acid levels were found to be a major factor in the CFS patients' regulation of plasma lipid concentration, but not in the controls. The alterations in the omega 6 fatty acids reflected the dysregulation of delta-6 desaturase activity and the supply of the prostaglandin precursor fatty acids. The changes observed in the CFS patients were consistent with a low-grade inflammation response. No evidence was found for changes in lipid homeostasis which reflected insulin resistance or changes in activities of delta-5 desaturase, elongase and stearoyl-CoA desaturase.
The increase in the plasma lipid concentrations in the CFS patients was associated with dysregulation of delta-6 desaturase activity and the omega 6 essential fatty acids, which were consistent with an inflammatory mediated event in the CFS patients.
THE DEVELOPMENT OF LABORATORY TESTS FOR INVESTIGATING CHRONIC FATIGUE AND PAIN DISORDERS [1998]
Dunstan RH, McGregor NR, Butt HL, Roberts TK, Zerbes M, Klineberg IJ
A diagnosis of chronic fatigue syndrome (CFS) requires the exclusion of other known fatigue related diseases. It also requires compliance with a clinical definition that is primarily characterised by unexplained, persistent or relapsing, chronic and debilitating fatigue lasting more than six consecutive months.
The patient set derived by this process is heterogeneous, and has proved very difficult to study clinically and scientifically. There is an urgent demand for a range of specialised tests to be developed to assist the clinicians in classifying the various types of chronic fatigue and chronic pain disorders. Data from studies involving analyses of plasma lipids and urinary excretion of metabolites together, with their associations with symptom presentation has been used to classify patients with similar metabolic profiles into subgroups.
These subgroups comprise relatively homogeneous sets of patients for investigation. Individual patients can also be matched to the metabolic profiles from the CPRU’s research to assign their altered homeostasis patterns to a particular CFS type.
In addition, the test data can provide insight into potential management strategies, as certain host responses can be determined, and nutrient deficiencies identified. Three main types of specialist tests are currently available. These are:
1. Urine test
Analyses of the metabolites in the urine can indicate whether the patient has a significantly altered pattern of excretion representing an altered homeostasis, and whether or not that patient can be assigned to a previously defined type of chronic fatigue disorder. The data provided can indicate whether fibrillar or non-fibrillar catabolism is active in the patient, and whether any anomalies identified in the excretion pattern provide objective evidence for gut, neurological, energy and urea cycle dysfunction. An indication of specific amino acid deficiencies can also be obtained.
2. Blood lipid test
Analyses of plasma lipid profiles can indicate anomalies in lipid homeostasis and enable the assignment of a patient to a previously defined type of chronic fatigue disorder. The test can detect deficiencies in essential fatty acid composition, detect dysregulation of essential fatty acid desaturation and elongation, and provide evidence for the presence of host responses to infectious agents.
3. Faecal Microbiology
Analyses of faecal aerobic and anaerobic micro-organisms in conjunction with faecal lipid analysis can provide extensive data on the functional integrity of the gastrointestinal tract. The data can indicate overgrowths of specific organisms such as Bacteroides and low numbers of critical organisms such as E. coli, Lactobacillus and Bifidobacterium species. This data can lead to developing strategies for reconstituting the bowel flora. The faecal lipid data can further indicate evidence for dysfunctional bowel flora. Evidence also be obtained for malabsorption, and the functional activities of the anaerobic organisms in their interconversions of bile and sterol products.
REFERENCES:
Holmes GP, Kaplan JE, Gantz NM, et al. Chronic fatigue syndrome: A working case definition. Ann. Intern. Med. 1988; 108:387-389.
Fukuda K, Straus SE, Hickie I, et al. The chronic fatigue syndrome: A comprehensive approach to its definition and study. Ann. Intern. Med. 1994; 121:953-959.
McGregor NR, Dunstan RH, Zerbes M, Butt HL, Roberts TK, Klineberg IJ. Preliminary determination of a molecular basis to chronic fatigue syndrome. Biochem. Molec. Med. 1996; 57: 73-80.
McGregor NR, Dunstan RH, Zerbes M, Butt HL, Roberts TK, Klineberg IJ. Preliminary determination of the association between symptom expression and urinary metabolites in subjects with chronic fatigue syndrome. Biochem. Molec. Med. 1996; 58:85-92.
McGregor NR, Dunstan RH, Butt HL, Zerbes M, Roberts TK, Klineberg IJ (1997). A preliminary assessment of the association of SCL-90-R psychological inventory responses with changes in urinary metabolites in patients with chronic fatigue syndrome. J Chronic Fatigue Syndrome 3:17-37
CHRONIC FACIAL MUSCLE PAIN AND DYSREGULATED CELLULAR PROTEOLYSIS [1998]
NR McGregor, RH Dunstan, HL Butt, TK Roberts, M Zerbes, IJ Klineberg.
Chronic muscle pain is a common condition for which there is no clearly identified aetiology, although its onset has been associated with infectious events, trauma and increasing life stresses. CFS patients have increased prevalence of facial muscle pain compared to the normal population.
Muscle pain has been associated with reductions in muscle protein and RNA concentrations. Thirty-five chronic facial muscle pain patients and 34 age and sex-matched control subjects were assessed for variations in urinary organic and amino acid excretion which could provide evidence of increased energy utilisation or proteolysis.
Compared to the controls, the MP patients had reductions in the excretion of leucine, which is an important amino acid for the regulation of proteolysis. The muscle pain patients also had increases in tyrosine excretion, which is a marker of proteolysis. (Proteolysis is the process by which proteins are broken down into amino acids and peptides.)
The visual analogue pain scale of average pain intensity (VAS) inversely correlated with the reduction in leucine excretion. The VAS score was positively related to the increased excretion of tyrosine, as well as the increased excretion of glutamic and aspartic acids (these are excitatory amino acids). No association was found between the VAS score and the fibrillar proteolysis marker, 3-methyl-histidine.
Increasing VAS scores were associated with an increased total amino acid output. This is consistent with the reductions in the serum amino acid levels observed in other studies and suggestive of a low grade aminoaciduria. The depletion of leucine may represent a significant anomaly in the regulation of proteolysis.
This data provides evidence to support the hypothesis that dysregulated non-fibrillar proteolysis occurs in chronic facial muscle pain patients. This has been found to be associated with increasing severity of chronic pain, as assessed by VAS scores.
ASSESSMENT OF LIPID HOMEOSTASIS IN SUDDEN AND GRADUAL ONSET CHRONIC FATIGUE SYNDROME PATIENTS [1998]
Neil McGregor, Hugh Dunstan, Mark Donohoe, Timothy Roberts, Henry Butt, Jennifer Watkins, Raymond Murdoch
A study was undertaken in 60 CFS patients and 39 age and sex-matched non-CFS control subjects. Plasma saponified lipid products were assessed using capillary gas chromatography - mass spectrometry to measure changes in plasma lipid in profiles. The major lipid anomalies observed in the CFS patients were reductions in trans-9-octadecenoate and in cholesterol.
Twenty of the CFS patients reported an “acute viral like” infection at onset and were assessed for Epstein-Barr virus (EBV) and Cytomegalovirus (CMV). The remaining 40 CFS patients reported a gradual onset. None of the sudden onset patients had immunological evidence of a current common viral infection. This observation was further substantiated by an absence of specific fatty acid anomalies that have been previously reported to be associated with single current acute or chronic viral infections.
The subgroup of CFS patients reporting a sudden onset had a different lipid profile compared to the controls, with the trans-9-octadecenoate : octadecanoate ratio as the primary inter-group difference. Those CFS patients reporting a gradual onset also had different lipid profiles compared to controls and the trans-9-octadecenoate : octadecanoate ratio was again the primary discriminant factor. Therefore, both sudden onset and gradual onset patients had the same fatty acid anomaly differentiating them from controls.
The sudden-onset CFS group lipid profile could be differentiated from the gradual onset group profile using the cis-9,12-octadecadienoate : cis-9-octadecanoate ratio. This ratio has been previously identified as a key post viral modification to lipid homeostasis.
Although the patient’s viral infective history and subsequent post viral modifications may play some role in determining the onset and progression of CFS, the primary lipid changes in CFS patients were related to other non virally induced lipid changes.
The differences in lipid homeostasis in the sudden and gradual onset groups provides a molecular basis for the heterogeneity observed in the CFS patients. This may have arisen from combinations of underlying genetic, dietary, immunological, environmental or pathogen altered responses.
LOW URINARY SERINE OUTPUT IS ASSOCIATED WITH AN ALTERED FAECAL MICROBIAL FLORA IN CHRONIC FATIGUE/PAIN PATIENTS
Butt H L, Dunstan R H, McGregor N R, Roberts T K, Harrison T L, Grainger JR
Serine is an important precursor of tryptophan and serotonin. Previous studies of patients with CFS demonstrated that serine was an important urinary metabolite discriminating CFS patients from control subjects. Serine synthesis requires both alanine and glycine as precursors, and also occurs as a result of microbial metabolism.
The aim of this study was to determine if faecal micro-organisms can produce serine, and how these organisms are quantitatively related to the total gastrointestinal microbial flora in CFS patients.
Three clinical faecal coliforms (Escherichia coli, Enterobacter cloacae and Proteus mirabilis) were incubated in defined broth and their metabolites were analysed by a gas chromatogram (GC-MS). Serine was produced by the three organisms at varying concentrations. Other amino acids including serine precursors (alanine and glycine), leucine, phenylalanine and succinic acid, were also detected. This suggested that the contribution of microbial metabolites by enteric organisms may be more important as previously thought.
Faecal samples from 27 CFS patients and 4 age and sex matched control subjects were studied. Quantitative bacteriology was performed on all samples. Seventeen (62.9%) CFS patients had a low % distribution of E.coli. In contrast, none of the four control subjects had a low % distribution of E.coli.
These studies showed that the low urinary excretion of serine in CFS patients reported in previous study was associated with a disturbed gastrointestinal microbial flora. Alteration in the aerobic microbial flora, particularly the gram negative enteric organisms, may change the external supply of serine and contribute to the increased symptoms expression of patients with CFS.
ASSESSMENT OF PLASMA LIPID HOMEOSTASIS IN RELATIONSHIP TO EPSTEIN BARR VIRUS ANTIBODY TITRES IN PATIENTS REPORTING SUDDEN ONSET CHRONIC FATIGUE [1998]
Neil McGregor, Hugh Dunstan, Mark Donohoe, Timothy Roberts, Henry Butt, Jennifer Watkins, Raymond Murdoch
A study was undertaken in 20 chronic fatigue syndrome patients who reported a sudden viral-like onset and 39 age and sex-matched control subjects. Plasma saponified lipid products were assessed using capillary gas chromatography -mass spectrometry to measure qualitative changes in plasma lipid profiles. Epstein Barr virus (EBV) IgM, IgG, early antigen (EBEA) and nuclear antigen (EBNA) were measured in the CFS patients.
Eighteen of the 20 sudden onset CFS patients had evidence of a past EBV infection. No patients had EBV IgM, 14 (70%) had positive EBV IgG titres and 14 (70%) were EBV nuclear antigen (EBNA) positive. Five patients (25%) were EBV early antigen (EBEA) positive. EBEA was associated with changes in cis-5,8,11,14-eicosatetraenoate, cholesterol, and other plasma lipid levels. This supported the observation that alterations in phospholipid hydrolysis and cholesterol metabolism are consistent with a cytokine response. (Cytokines are chemicals that are normally released as a result of the immune system's response to an infection).
These cytokine associated lipid changes, as well as changes associated with EBNA and EBV IgG, were not associated with the lipid alterations distinguishing CFS patients from control subjects reported previously. The lipid changes differentiating CFS patients from control subjects may be related to combinations of genetic, dietary, immunological, environmental or pathogen altered responses. © Bioscreen, CPRU, S Ashton and F Bartosy 1999 - 2001
N McGregor, H Dunstan, S Niblet, K King, H Butt, T Harrison, W Taylor, T Roberts, I Klineberg, V Clifton-Bligh, G Fulcher, J Dunsmore, L Hoskins
Ninety-eight CFS patients and 81 age and sex matched controls were assessed for the changes in symptoms, SCL-90-R psychological responses, and blood cell parameters. They were also examined for variations in blood biochemistry and the urinary excretion of organic and amino acids in relationship to their dental amalgam status. There were no variations detected due to the presence or quantities of dental amalgams in the number of subjects who reported improvement following removal of their amalgams, between the CFS patients and the control subjects.
No amalgam parameter was associated with CFS. There was no association between increasing numbers of amalgams or amalgam removal and any alterations in the clinical or patient-reported symptoms (prevalence, severity), SCL-90-R psychological responses, blood cell parameters, blood chemistry or urinary excretion of organic and amino acids in either the CFS patients or the control subjects.
Those subjects who perceived improvements in their symptoms following amalgam removal did not have any detectable difference in clinical or patient-reported (prevalence, severity) symptoms, SCL-90-R psychological responses, blood cell parameters, blood chemistry or urinary excretion of organic and amino acids.
Therefore, no differences in symptom expression or pathology could be detected in either CFS patients or control subjects for any amalgam parameters. No evidence was found that could support the removal of dental amalgam fillings in either CFS patients or control subjects for relieving poly-symptomatic presentation.
ALTERATIONS IN URINARY AMINO AND ORGANIC ACID EXCRETION IN PATIENTS WITH CHRONIC FATIGUE SYNDROME [1998]
Suzanne Niblett, Leigh Hoskin, Hugh Dunstan, Phillip Clifton-Bligh, Neil McGregor, Timothy Roberts, Greg Fulcher, Henry Butt, Katrina King, Julie Dunsmore, ML Neary, Tracey Harrison, Warren Taylor
One hundred CDC-defined CFS patients (27 males, 73 females) and 83 healthy control patients (23 males, 60 females) were recruited for a large collaborative study designed to:
1. Assess alterations in metabolism and homeostasis associated with CFS.
2. Relate these changes to physical and neuropsychological symptom expression and changes in nose and throat microbiota.
3. Evaluate the homogeneity of a CDC-defined CFS group.
All subjects were clinically reviewed and asked to complete questionnaires addressing subject demographics, medical history, presentation of signs, symptoms and sensitivities, psychological status and cognitive function.
Preliminary statistical analyses revealed significant differences in urinary metabolite excretion profiles in CFS patients compared to healthy control subjects. These results implied that changes in metabolism and homeostasis were associated with CFS. The urinary profiles of CFS subjects had increases in tyrosine, 3-methylhistidine, reductions in succinic acid, asparagine, phenylalanine, and hippuric acid and changes in several unidentified compounds. Reductions in the excretion rate of alanine, valine, leucine, proline and S-methylcysteine were also associated with CFS. This data was suggestive of alterations in protein and energy metabolism and implied changes in gut bacteria.
Male and female CFS and control subjects could be separated into four groups on the basis of their urinary metabolite profiles suggesting that the profiles were distinctive. The data suggested that gender specific anomalies occur in CFS patients compared to controls. Age was also significantly correlated with changes in the multivariate urinary metabolite excretion profiles in this study. When the groups were further divided into under/over 25 year old age groups, the CFS control group differences were again distinct. These findings suggest that the pathophysiology of CFS is different in females compared to males and that these differences are themselves influenced by age.
It was concluded that alterations in urine metabolite excretion by CFS patients were indicative of changes in metabolism and homeostasis in CFS patients. Preliminary evaluation of the CDC-defined CFS group homogeneity found that sex and age contributed to heterogeneity in the CFS group and suggested that these factors should be considered in future studies of CFS
patients.
CHLORINATED PESTICIDES AND CHRONIC FATIGUE RELATED ILLNESS
Hugh Dunstan, Neil McGregor, Warren Taylor, Mark Donohoe, Timothy Roberts, Henry Butt, Raymond Murdoch, Jennifer Watkins
Chronic fatigue syndrome is a disorder characterised by chronic fatigue/pain symptoms, and the definition includes many exclusionary criteria (ref. 1).
Hexachlorobenzene (HCB) and dichlorodiphenyltrichloroethane (DDT) are chlorinated hydrocarbon pesticides, which are molecules that accumulate in the membranes of all cells in the body. This represents a substantial distribution of pesticides around the various body organs. The integrity of the membranes in which these chemicals accumulate is essential to proper function of the cells and communication between cells. The organochlorines can interact with the membranes to alter fluidity properties and to disrupt membrane protein function.
Laboratory findings by the CPRU (refs. 2 and 3) have indicated that CFS patients can have significantly higher levels of serum DDT compared to control subjects. In addition, DDE and HCB levels correlated with changes in blood cell parameters such as red cell distribution width and haemoglobin content.
Exploration of the early literature revealed interesting associations of pesticide usage and viral activities, as well as an association of DDT with substantial alterations in metabolic homeostasis.
References:
1. Holmes, G.P., Kaplan, J.E., Gantz, N.M. et al (1988) Chronic fatigue syndrome: a working case definition. Ann Intern. Med. 108, 387- 389.
2. Dunstan RH, Donohoe M, Taylor W, et al. A preliminary investigation of chlorinated hydrocarbons and chronic fatigue syndrome. Med J Aus 1995; 163:294-297.
3. Dunstan RH, Roberts TK, Donohoe M, McGregor NR, Hope D, Taylor WG, Watkins JA, Murdoch RN & Butt H (1996) Bioaccumulated chlorinated hydrocarbons and red/white blood cell parameters. Biochem. Molec.Med. 58:77-84.
CHRONIC PAIN AND PROTEIN TURNOVER IN POLYSYMPTOMATIC PATIENTS
Neil McGregor, Hugh Dunstan, Henry Butt, Timothy Roberts, Iven Klineberg, Phillip Clifton-Bligh, Greg Fulcher, Julie Dunemore, Leigh Hoskin
Chronic pain is one of the major reasons for seeking medical treatment, however the mechanisms by which chronic pain occurs are very poorly understood. Onset of chronic pain has been associated with infectious events, trauma and increasing life stresses, each of which may induce an increased host energy demand. These factors suggest that dysregulation of protein turnover is associated with chronic pain.
The data from three studies assessed the potential mechanisms of chronic pain seen in chronic polysymptomatic patients. The results show that alteration in the tyrosine:leucine ratio (a marker of the relationship between proteolysis and protein synthesis) is strongly associated with chronic muscle pain.
Enhanced proteolysis is associated with diuresis and an increase in the secretion of excitatory amino acids, such as glutamic and aspartic acids. Prolonged muscle pain is associated with increases in the urea-markers of tissue damage. This data supports an alteration in nitric oxide production and enhanced NMDA activation, two factors essential for the initiation of spinal column hyperalgesia.
The inhibition of these responses is mediated by noradrenaline, and may explain why certain tricyclic antidepressants may be useful in management of the pain response. Understanding of the mechanism behind chronic pain is essential for the development of appropriate therapies.
BIOCHEMICAL ABNORMALITIES IN CHRONIC FATIGUE SYNDROME
Phillip Clifton-Bligh, Suzanne Niblett, Leigh Hoskin, Hugh Dunstan, Greg Fulcher, Neil McGregor, Julie Dunsmore, Timothy Roberts, Henry Butt, Katrina King, Iven Klineberg
The overnight urine excretion rate of a number of amino acids and organic acids was measured in 100 patients with chronic fatigue syndrome, and in 83 age and sex matched control subjects.
The excretion rate of the metabolites was measured in units/minute. The most striking differences between patients with chronic fatigue syndrome and controls was a reduction in urine asparagine (p<0.0001) and a reduction in urine succinic acid (p<0.0003) in patients with CFS.
Urinary tyrosine (p<0.04) and urinary 3-methyl histidine (p<0.03) were significantly increased in patients with chronic fatigue syndrome. The urinary excretion of tyrosine, a protein catabolism marker, was associated with symptoms of fatigue, muscle pain, lymph node pain and cognitive disturbance.
The overnight excretion of succinic acid was correlated with the fasting plasma glucose (p<0.002). It is not known at present whether urine excretion rates of succinic acid and asparagine are correlated with serum concentrations of these metabolites. It is also not clear whether renal tubular function independently influences the urine excretion rates, or if renal tubular function is itself altered in chronic fatigue syndrome.
It is possible that urine asparagine excretion rates are low because of reduced entry of asparagine into the circulation, and therefore into the urine. If the rate of entry of asparagine into the circulation is reduced, this could come about because of more rapid metabolism of asparagine in cells, or because total body stores of asparagine are depleted. Further work is necessary to define these issues.
FAECAL MICROBIAL GROWTH INHIBITION IN CHRONIC FATIGUE/PAIN PATIENTS
Butt HL, Dunstan RH, McGregor NR, Roberts TK, Harrison TL, Grainger JR
Chronic fatigue/pain patients have reported frequent gastrointestinal symptoms (irritable bowel, constipation, abdominal pain, diarrhoea, nausea) suggesting the possibility of an altered microbial flora in the gastrointestinal tract. The aim of this investigation was to determine if there was a change in the microbial intestinal flora of patients with CFS, and what causative factors may be implicated for these changes. Faecal samples from 27 polysymptomatic CFS patients and four age and sex matched control subjects were studied. © Bioscreen, CPRU, S Ashton and F Bartosy 1999 - 2001
Quantitative bacteriology of three aerobes (Escherichia coli, Klebsiella/Enterobacter group, and Enterococcus faecalis) and three anaerobes (Bacteroides spp., Bifidobacterium spp. and Lactobacillus spp.) was performed on all samples. The incidence of CFS patients with faecal Escherichia coli greater than the percentage mean of control subjects was significantly different to that of the Bacteroides spp. (7 vs 21 respectively, p=0.0001) suggesting the possibility of an antimicrobial interaction among bacterial species.
Metabolites of Bacteroides spp. (lactic, propionic and butyric acids) were examined for growth inhibitory properties against three clinical faecal coliform (Escherichia coli, Klebsiella and Proteus mirabilis). All three coliforms were susceptible to the three short chain fatty acids at pH 3-4. At pH 6-7 the three coliforms were not susceptible to lactic acid, partially to propionic acid, and fully to butyric acid.
This study found marked alteration of the coliforms and lactic acid bacteria in the faecal microbial flora of CFS patients. It was concluded that these changes may be related to inter-microbial activities between organisms.
ANTI-NUCLEAR ANTIBODIES AND CFS
NR McGregor, RH Dunstan, S Niblet, K King, HL Butt, T Harrison, W Taylor, TK Roberts, IJ Klineberg, P Clifton-Bligh, G Fulcher, J Dunsmore, L Hoskins
Ninety-eight CFS patients and 81 age and sex matched controls were assessed for the changes in symptoms, SCL-90-R psychological responses, blood cell parameters, blood biochemistry and the urinary excretion of organic and amino acids in relationship to their anti-nuclear antibody (ANA) status. CFS patients were divided into ANA positive and ANA negative groups for comparison.
The fatigue responses in the ANA positive patients were more severe than those observed in the remaining negative group. The fatigue responses were associated with alterations in red blood cell parameters, which were consistent with an increased compensated haemolysis reaction. Increasing ANA titres were associated with alterations in haemoglobin and red cell volume as well as increases in iron levels and reductions in sodium levels. No change in any immune cell parameter was found. These responses were quite different from those noted in the CFS ANA negative patients.
The ANA positive patients had an increase in depression scores, which were found to be associated with alterations in red blood cell parameters and increased levels of bilirubin and the bilirubin:albumin ratio. This indicates a potential chemical neurotoxicity and not a psychological depressive state.
This data was consistent with an auto-immune associated alteration in red cell haemolysis, which has been previously associated with the presentation of fatigue in mild hepatobillary disorders or porphyrin disorders. These types of fatigue/pain illnesses need to be diagnosed and excluded under the current CFS definition.
TOXIC COAGULASE NEGATIVE STAPHYLOCOCCI ARE ASSOCIATED WITH CHANGES IN URINARY ORGANIC AND AMINO ACID EXCRETION IN CHRONIC FACIAL MUSCLE PAIN PATIENTS
NR McGregor, HL Butt, RH Dunstan, TK Roberts, M Zerbes, IJ Klineberg
This study examined 35 patients with chronic facial muscle pain and 34 age and sex matched control subjects. The purpose of this investigation was to assess how staphylococcal bacteria can influence symptoms and changes in urinary excretion of amino and organic acids.
The muscle pain patients had an increase in the carriage of toxic coagulase negative species (TCoNS) which produced either delta toxin or both delta and “horse” toxins. The carriage of TCoNS was associated with increases in pain severity, irritable bowel symptoms, palpitations, muscle fatigue and recurrent low-grade fever consistent with the symptom profile of fibromyalgia.
The carriage of TCoNS was inversely related to the reduced excretion of leucine, which is an important modulator of proteolysis. The carriage of TCoNS was positively associated with increased excretions of tyrosine and 3-methylhistidine.
Other anomalies were also associated with the carriage of TCoNS. These included increased excretions of ornithine, aconitic acid, hydroxyproline, proline and aspartic acid. In addition, there was an increased excretion of total amino acids suggestive of a low-grade aminoaciduria.
The findings of this study indicated that toxic coagulase negative staphylococcal bacteria were strongly correlated with anomalies in proteolysis and other aspects of the metabolism. This implies that these organisms may have a major role in the aetiology of chronic muscle pain.
I Buttfield, H Butt, H Dunstan, N McGregor, T Roberts
Chronic fatigue syndrome (CFS) has had a very controversial history. The existence of this syndrome continues to be questioned by some, but now we have overwhelming evidence supporting the belief that this is an "organic" disorder. Recent research has suggested that this disorder may be related to cell wall dysfunction. The reason is not understood as yet, but new research continues to try to explain this polysymptomatic disorder. © Bioscreen, CPRU, S Ashton and F Bartosy 1999 - 2001
Given the controversy surrounding the diagnosis, it is not surprising that treatment has become confused and emotional. Many treatments continue to be tried and some partial successes have been found.
Treatments fall into formats such as "medically acceptable" therapies that attempt to treat some symptoms, to a range of unconventional remedies that claim to treat fatigue and improve overall health. Many treatments have tended to focus upon vitamins and health foods in the belief that the fatigue is due to some changes in the "milieu interna".
Such therapies are unsuccessful in part because the disease has causes that few have expected. Some of these therapies have had clinical trial data to validate them, although none have had a real curative purpose. The best that could be hoped for, was, a slight improvement in function and the doctor has not always been helpful even with that. There is an increasing public demand for more effective therapies and proper support, not only from doctors but also from skilled medical therapists such as dietitians.
With the advent of the discovery of the "molecular biology" of CFS, there is real hope of developing effective treatment strategies for this disorder. The CPRU, and now Bioscreen, continues to be heavily involved in this promising new area.
HOMEOSTATIC HETEROGENEITY IN CFS PATIENTS [1998]
Dunstan RH, McGregor NR, Butt HL, Roberts TK, Zerbes M, Klineberg IJ
The original definition of CFS published by the American Centres for Disease Control (CDC) was initially conceived to treat CFS as a potentially infectious disease, as evidenced by the large number of well-documented outbreaks.
A diagnosis of chronic fatigue syndrome requires the exclusion of other known fatigue-related diseases and requires compliance with a clinical definition which is primarily characterised by unexplained, persistent or relapsing, chronic and debilitating fatigue lasting six or more months. The patient group derived by this process is heterogeneous in its presentation and has proved very difficult to study clinically and scientifically.
Analyses of the excretion of metabolites in the urine have indicated that CFS patients have a significantly altered pattern of excretion suggestive of an altered homeostasis. Increases in CFSUM1, Beta-alanine and tyrosine (a marker for non-fibrillar proteolysis), and decreases in serine, alanine and succinic acid were observed.
Elevations in CFSUM1 and beta-alanine were positively associated with increased symptom incidence and severity as well as musculo-skeletal symptoms and gastrointestinal symptoms. Conversely, reduced output of serine was associated with increasing severity of neurological dysfunction. This data provides evidence of a relationship between alterations in homeostasis and symptom expression.
Analyses of the CFS biochemical data by cluster analysis and multivariate techniques indicated that different types of CFS patients could be characterised on the basis of similarities in the metabolite profiles. The urine profiles can therefore be used to classify patients with similar excretion patterns to give relatively uniform sets of patients for investigation.
In a similar manner, the plasma fatty acid profile can be used to differentiate CFS patients from control subjects. The CFS patients can be further characterised by clustering techniques to align patients with similar fatty acid homeostasis.
The capacity to classify CFS patients into subgroups using objectively derived measurements represents a major advance in CFS research. This is because it provides the capacity to develop treatment regimes based on objectively measured parameters, and minimises the confounding effects of heterogeneity in future research programs.
References:
1. Holmes GP, Kaplan JE, Gantz NM, et al. Chronic fatigue syndrome: A working case definition. Ann Intern Med 1988; 108:387-389.
2. Fukuda K, Straus SE, Hickie I, et al. The chronic fatigue syndrome: A comprehensive approach to its definition and study. Ann Intern Med 1994; 121:953-959.
3. McGregor NR, Dunstan RH, Zerbes M, Butt HL, Roberts TK, Klineberg IJ. Preliminary determination of a molecular basis to chronic fatigue syndrome. Biochem Molecul Med 1996; 57: 73-80.
4. McGregor NR, Dunstan RH, Zerbes M, Butt HL, Roberts TK, Klineberg IJ. Preliminary determination of the association between symptom expression and urinary metabolites in subjects with chronic fatigue syndrome. Biochem Molec Med 1996; 58:85-92.
© CPRU and F Bartosy 1999
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