Sonya Marshall-Gradisnik

Immunological abnormalities as potential biomarkers in Chronic Fatigue Syndrome/Myalgic Encephalomyelitis

BackgroundChronic Fatigue Syndrome/Myalgic Encephalomyelitis (CFS/ME) is characterised by severe prolonged fatigue, and decreases in cognition and other physiological functions, resulting in severe loss of quality of life, difficult clinical management and high costs to the health care system. To date there is no proven pathomechanism to satisfactorily explain this disorder. Studies have identified abnormalities in immune function but these data are inconsistent. We investigated the profile of markers of immune function (including novel markers) in CFS/ME patients.MethodsWe included 95 CFS/ME patients and 50 healthy controls. All participants were assessed on natural killer (NK) and CD8+T cell cytotoxic activities, Th1 and Th2 cytokine profile of CD4+T cells, expression of vasoactive intestinal peptide receptor 2 (VPACR2), levels of NK phenotypes (CD56bright and CD56dim) and regulatory T cells expressing FoxP3 transcription factor.ResultsCompared to healthy individuals, CFS/ME patients displayed significant increases in IL-10, IFN-γ, TNF-α, CD4+CD25+ T cells, FoxP3 and VPACR2 expression. Cytotoxic activity of NK and CD8+T cells and NK phenotypes, in particular the CD56bright NK cells were significantly decreased in CFS/ME patients. Additionally granzyme A and granzyme K expression were reduced while expression levels of perforin were significantly increased in the CFS/ME population relative to the control population. These data suggest significant dysregulation of the immune system in CFS/ME patients.ConclusionsOur study found immunological abnormalities which may serve as biomarkers in CFS/ME patients with potential for an application as a diagnostic tool.

Longitudinal investigation of natural killer cells and cytokines in chronic fatigue syndrome/myalgic encephalomyelitis

BackgroundChronic Fatigue Syndrome/Myalgic Encephalomyelitis (CFS/ME) is an etiologically unexplained disorder characterised by irregularities in various aspects of the immunological function. Presently, it is unknown whether these immunological changes remain consistent over time. This study investigates Natural Killer (NK) cell cytotoxic activity, NK cell subsets (CD56brightCD16- and CD56dimCD16+) and cytokines, over the course of a12 month period in patients with CFS/ME.MethodsThe participants in the study comprised 65 (47.2 ± 11.5 years) CFS/ME participants and 21 (45.2 ±9.3 years) non-fatigued controls. Flow cytometry protocols were used to assess NK subsets and NK cytotoxic activity at various time points that included baseline (T1), 6 (T2) and 12 months (T3). Cytokine secretions were measured following mitogenic stimulation of peripheral blood mononuclear cells.ResultsNK cytotoxic activity was significantly decreased in the CFS/ME patients at T1, T2 and T3 compared to the non-fatigued group. Additionally, in comparison to the non-fatigued controls, the CFS/ME group had significantly lower numbers of CD56brightCD16- NK cells at both T1 and T2. Interestingly, following mitogenic stimulation, cytokine secretion revealed significant increases in IL-10, IFN-γ and TNF-α at T1 in the CFS/ME group. A significant decrease was observed at T2 in the CFS/ME group for IL-10 and IL-17A while at T3, IL-2 was increased in the CFS/ME group in comparison to the non-fatigued controls. Overall cytotoxic activity was significantly decreased at T3 compared to T1 and T2. CD56brightCD16- NK cells were much lower at T2 compared to T1 and T3. IL-10 and IL-17A secretion was elevated at T2 in comparison to T1 and T3.ConclusionThese results confirm decreases in immune function in CFS/ME patients, suggesting an increased susceptibility to viral and other infections. Furthermore, NK cytotoxic activity may be a suitable biomarker for diagnosing CFS/ME as it was consistently decreased during the course of the 12 months study.

Effects of estrogen on the mechanical behavior of the human Achilles tendon in vivo

The purpose of this study was to elucidate the effect of normal fluctuating [non-monophasic oral contraceptive pill (MOCP) users] and low, consistent (MOCP users) endogenous plasma estrogen levels on the strain behavior of the Achilles tendon in vivo. Twenty women (age 28.0 +/- 4.2 yr, height 1.67 +/- 0.07 m, mass 61.6 +/- 6.8 kg) who had been using the MOCP for at least 12 mo together with 20 matched women who were non-MOCP users (age 31.9 +/- 7.3 yr, height 1.63 +/- 0.05 m, mass 62.5 +/- 5.9 kg) participated in this study. Non-MOCP users were tested at the time of lowest (menstruation) and highest (approximately same as ovulation) estrogen, whereas MOCP users, who exhibited constant and attenuated endogenous estrogen levels, were tested at day 1 and day 14 of their cycle. At each test session, maximal isometric plantarflexion efforts were performed on a calf-raise apparatus while synchronous real-time ultrasonography of the triceps surae aponeurosis was recorded. Achilles tendon strain (%) was calculated by dividing tendon displacement during plantarflexion by resting tendon length. Repeated-measures ANOVA revealed a significant (P < 0.05) main effect of subject group with significantly lower Achilles strain (25.5%) in the MOCP users compared with the non-MOCP users. In conclusion, acute fluctuations in plasma estrogen across the menstrual cycle in non-MOCP users did not alter the strain behavior of the Achilles tendon. Conversely, long-term exposure to attenuated estrogen in MOCP users resulted in a decrease in Achilles tendon strain, which is thought to be attributed to the effects of endogenous estrogen on collagen synthesis. These findings have a number of important functional and clinical implications.

Immune and hemorheological changes in Chronic Fatigue Syndrome

BackgroundChronic Fatigue Syndrome (CFS) is a multifactorial disorder that affects various physiological systems including immune and neurological systems. The immune system has been substantially examined in CFS with equivocal results, however, little is known about the role of neutrophils and natural killer (NK) phenotypes in the pathomechanism of this disorder. Additionally the role of erythrocyte rheological characteristics in CFS has not been fully expounded. The objective of this present study was to determine deficiencies in lymphocyte function and erythrocyte rheology in CFS patients.MethodsFlow cytometric measurements were performed for neutrophil function, lymphocyte numbers, NK phenotypes (CD56dimCD16+ and CD56brightCD16-) and NK cytotoxic activity. Erythrocyte aggregation, deformability and fibrinogen levels were also assessed.ResultsCFS patients (n = 10) had significant decreases in neutrophil respiratory burst, NK cytotoxic activity and CD56brightCD16- NK phenotypes in comparison to healthy controls (n = 10). However, hemorheological characteristic, aggregation, deformability, fibrinogen, lymphocyte numbers and CD56dimCD16+ NK cells were similar between the two groups.ConclusionThese results indicate immune dysfunction as potential contributors to the mechanism of CFS, as indicated by decreases in neutrophil respiratory burst, NK cell activity and NK phenotypes. Thus, immune cell function and phenotypes may be important diagnostic markers for CFS. The absence of rheological changes may indicate no abnormalities in erythrocytes of CFS patients.

The effect of five weeks of Tribulus terrestris supplementation on muscle strength and body composition during preseason training in elite rugby league players.

Tribulus terrestris is an herbal nutritional supplement that is promoted to produce large gains in strength and lean muscle mass in 5–28 days (15, 18). Although some manufacturers claim T. terrestris will not lead to a positive drug test, others have suggested that T. terrestris may increase the urinary testosterone/epitestosterone (T/E) ratio, which may place athletes at risk of a positive drug test. The purpose of the study was to determine the effect of T. terrestris on strength, fat free mass, and the urinary T/E ratio during 5 weeks of preseason training in elite rugby league players. Twenty-two Australian elite male rugby league players (mean ± SD; age = 19.8 ± 2.9 years; weight = 88.0 ± 9.5 kg) were match-paired and randomly assigned in a double-blind manner to either a T. terrestris (n = 11) or placebo (n = 11) group. All subjects performed structured heavy resistance training as part of the clubs preseason preparations. A T. terrestris extract (450 mg·d-1) or placebo capsules were consumed once daily for 5 weeks. Muscular strength, body composition, and the urinary T/E ratio were monitored prior to and after supplementation. After 5 weeks of training, strength and fat free mass increased significantly without any between-group differences. No between-group differences were noted in the urinary T/E ratio. It was concluded that T. terrestris did not produce the large gains in strength or lean muscle mass that many manufacturers claim can be experienced within 5–28 days. Furthermore, T. terrestris did not alter the urinary T/E ratio and would not place an athlete at risk of testing positive based on the World Anti-Doping Agencys urinary T/E ratio limit of 4:1.

Effect of exercise intensity on bone density, strength, and calcium turnover in older women

PURPOSE This study examined the effects of 24 wk of high intensity strength training or low intensity walking on lumbar bone mineral density (BMD), muscular strength, and calcium turnover in Australian women either taking hormone replacement therapy (HRT) or not taking HRT. METHODS A subject pool of 64 women between 45-65 yr and randomly allocated into weights (N = 21), walking (N = 20), weightsHRT (N = 14), and walkingHRT (N = 9) groups completed this study. All subjects trained twice weekly in either a 50-min walking or weight-training program (60-90% IRM). Measurements included maximal isometric knee strength, IRM bench press, IRM squat, isokinetic back strength, lumbar (L2-L4) BMD, serum osteocalcin, and urinary deoxypyridinoline crosslinks (Dpd). RESULTS No significant group differences in BMD were evident at the completion of training. However, a significant (P < 0.05) within group change was apparent for the walking group since BMD decreased 1.3% below baseline testing. Osteocalcin levels increased significantly (P < 0.05) in the walking (22%) group. Maximal bench press and squat strength improved significantly (P < 0.05) in the weights (25.8% and 37.7%) and weightsHRT (25.4% and 35.7%) groups. The weights group also increased significantly (P < 0.05) in isokinetic back strength (22.2%). CONCLUSION It was concluded that short-term high intensity resistance training provides an effective means for increasing muscular strength in women between 45 and 65 yr. The training effects on lumbar BMD were not apparent in the present study.

The prevalence of chronic fatigue syndrome/myalgic encephalomyelitis: a meta-analysis

Purpose To perform a meta-analysis to examine variability among prevalence estimates for CFS/ME, according to the method of assessment used. Methods Databases were systematically searched for studies on CFS/ME prevalence in adults that applied the 1994 Centers for Disease Control (CDC) case definition.1 Estimates were categorized into two methods of assessment: self-reporting of symptoms versus clinical assessment of symptoms. Meta-analysis was performed to pool prevalences by assessment using random effects modeling. This was stratified by sample setting (community or primary care) and heterogeneity was examined using the I2 statistic. Results Of 216 records found, 14 studies were considered suitable for inclusion. The pooled prevalence for self-reporting assessment was 3.28% (95% CI: 2.24–4.33) and 0.76% (95% CI: 0.23–1.29) for clinical assessment. High variability was observed among self-reported estimates, while clinically assessed estimates showed greater consistency. Conclusion The observed heterogeneity in CFS/ME prevalence may be due to differences in method of assessment. Stakeholders should be cautious of prevalence determined by the self-reporting of symptoms alone. The 1994 CDC case definition appeared to be the most reliable clinical assessment tool available at the time of these studies. Improving clinical case definitions and their adoption internationally will enable better comparisons of findings and inform health systems about the true burden of CFS/ME.

Role of adaptive and innate immune cells in chronic fatigue syndrome/myalgic encephalomyelitis

Perturbations in immune processes are a hallmark of a number of autoimmune and inflammatory disorders. Chronic fatigue syndrome/myalgic encephalomyelitis (CFS/ME) is an inflammatory disorder with possible autoimmune correlates, characterized by reduced NK cell activity, elevations in regulatory T cells (Tregs) and dysregulation in cytokine levels. The purpose of this article is to examine innate and adaptive immune cell phenotypes and functional characteristics that have not been previously examined in CFS/ME patients. Thirty patients with CFS/ME and 25 non-fatigued controls were recruited for this study. Whole blood samples were collected from all participants for the assessment of cell phenotypes, functional properties, receptors, adhesion molecules, antigens and intracellular proteins using flow cytometric protocols. The cells investigated included NK cells, dendritic cells, neutrophils, B cells, T cells, γδT cells and Tregs. Significant changes were observed in B-cell subsets, Tregs, CD4(+)CD73(+)CD39(+) T cells, cytotoxic activity, granzyme B, neutrophil antigens, TNF-α and IFN-γ in the CFS/ME patients in comparison with the non-fatigued controls. Alterations in B cells, Tregs, NK cells and neutrophils suggest significant impairments in immune regulation in CFS/ME and these may have similarities to a number of autoimmune disorders.

The effect of short-term use of testosterone enanthate on muscular strength and power in healthy young men

Use of testosterone enanthate has been shown to significantly increase strength within 6–12 weeks of administration (2, 9), however, it is unclear if the ergogenic benefits are evident in less than 6 weeks. Testosterone enanthate is classified as a prohibited substance by the World Anti- Doping Agency (WADA) and its use may be detected by way of the urinary testosterone/epitestosterone (T/E) ratio (16). The two objectives of this study were to establish (a) if injection of 3.5 mg·kg-1 testosterone enanthate once per week could increase muscular strength and cycle sprint performance in 3–6 weeks; and (b) if the WADA-imposed urinary T/E ratio of 4:1 could identify all subjects being administered 3.5 mg·kg-1 testosterone enanthate. Sixteen healthy young men were match-paired and were assigned randomly in a double-blind manner to either a testosterone enanthate or a placebo group. All subjects performed a structured heavy resistance training program while receiving either testosterone enanthate (3.5 mg·kg-1) or saline injections once weekly for 6 weeks. One repetition maximum (1RM) strength measures and 10-second cycle sprint performance were monitored at the pre (week 0), mid (week 3), and post (week 6) time points. Body mass and the urinary T/E ratio were measured at the pre (week 0) and post (week 6) time points. When compared with baseline (pre), 1RM bench press strength and total work during the cycle sprint increased significantly at week 3 (p < 0.01) and week 6 (p < 0.01) in the testosterone enanthate group, but not in the placebo group. Body mass at week 6 was significantly greater than at baseline in the testosterone enanthate group (p < 0.01), but not in the placebo group. Despite the clear ergogenic effects of testosterone enanthate in as little as 3 weeks, 4 of the 9 subjects in the testosterone enanthate group (−44%) did not test positive to testosterone under current WADA urinary T/E ratio criteria.

Cytotoxic lymphocyte microRNAs as prospective biomarkers for Chronic Fatigue Syndrome/Myalgic Encephalomyelitis

BACKGROUND Immune dysfunction associated with a disease often has a molecular basis. A novel group of molecules known as microRNAs (miRNAs) have been associated with suppression of translational processes involved in cellular development and proliferation, protein secretion, apoptosis, immune function and inflammatory processes. MicroRNAs may be implicated in Chronic Fatigue Syndrome/Myalgic Encephalomyelitis (CFS/ME), where immune function is impaired. The objective of this study was to determine the association between miRNAs in cytotoxic cells and CFS/ME. METHODS Natural Killer (NK) and CD8(+)T cells were preferentially isolated from peripheral blood mononuclear cells from all participants (CFS/ME, n=28; mean age=41.8±9.6 years and controls, n=28; mean age=45.3±11.7 years), via negative cell enrichment. Following total RNA extraction and subsequent synthesis of cDNA, reverse transcriptase-quantitative polymerase chain reaction (RT-qPCR) was used to determine the expression levels of nineteen miRNAs. RESULTS There was a significant reduction in the expression levels of miR-21, in both the NK and CD8(+)T cells in the CFS/ME sufferers. Additionally, the expression of miR-17-5p, miR-10a, miR-103, miR-152, miR-146a, miR-106, miR-223 and miR-191 was significantly decreased in NK cells of CFS/ME patients in comparison to the non-fatigued controls. LIMITATIONS The results from these investigations are not yet transferable into the clinical setting, further validatory studies are now required. CONCLUSIONS Collectively these miRNAs have been associated with apoptosis, cell cycle, development and immune function. Changes in miRNAs in cytotoxic cells may reduce the functional capacity of these cells and disrupt effective cytotoxic activity along with other immune functions in CFS/ME patients.