From Discovery to Application

By Dane Cook, PhD
University of Wisconsin-Madison

Last summer, CFIDS Association CEO Kim McCleary provided an excellent overview of the research being conducted by Drs. Alan and Kathy Light at the University of Utah using exercise to study gene expression in chronic fatigue syndrome (CFS) patients. Two intriguing questions were posted on Research1st in response to the overview:

1. How do these findings fit with the study on a proteomic signature in spinal fluid and is there any overlap between the gene expression in this study and any of the proteins in the spinal fluid study?
2. How large of a study needs to be done in order to validate the Lights’ findings?

First allow me to provide a very brief summary of the two studies referenced, one from the Light team and the other from Dr. Steven Schutzer’s team at the University of Medicine and Dentistry of New Jersey (UMDNJ):

Light, Light and colleagues addressed a critical aspect of CFS — post-exertion malaise (PEM). Thus, their study examined a clinical aspect of the disease. Based on their previous work examining receptors for adenosine tri-phosphate, hydrogen ions and vanilloid/heat receptors on mouse sensory neurons, they took a novel approach and examined messenger RNA (mRNA) for these same receptors on white blood cells of CFS patients, fibromyalgia patients and controls. Measures were taken both prior to and following moderately intense exercise. Their results indicated that CFS patients (both with and without FM) did not differ from controls at baseline; however, patients with FM only did differ at baseline. CFS patients showed significantly enhanced expression for receptors involved in cardiovascular, immune and sensory regulation after the exercise challenge; however, FM patients did not show enhanced gene expression post exercise. In the CFS patients, increases in gene expression were significantly and positively related to symptoms of pain and fatigue. This study emphasized the importance of fatigue provocation in CFS research and provided a potential objective biological mechanism in support of the patients’ subjective experience and as potential targets for therapeutic intervention trials.

In February of 2011, Schutzer and colleagues at UMDNJ, Pacific Northwest National Laboratory and Columbia University reported results from another ground-breaking study using proteomics to study cerebral spinal fluid (CSF) levels of proteins in CFS patients, patients with neurologic post-treatment Lyme disease syndrome (nPTLS) and healthy controls. This study was a cross-sectional or “snapshot” look at proteins in the central nervous system and was not intended to explain any particular aspect of illness. The data indicated 738 unique proteins for CFS patients (no overlap with nPTLS or controls), 692 unique proteins for nPTLS and 724 unique proteins for healthy controls. The authors made the following conclusions:

• nPTLS and CFS have distinguishing spinal fluid protein complements
• Each condition has a number of spinal fluid proteins that can be useful in providing candidates for future validation studies and insights on the respective mechanisms of pathogenesis
• Distinguishing nPTLS and CFS permits more focused study of each condition and can lead to novel diagnostics and therapeutic interventions.

The short answer to the first question is that the Light and Schutzer studies actually have very little overlap. They differ in sample origin – peripheral blood and leukocytes for Light and spinal fluid for Schutzer. They differ in the primary outcome variable – gene expression for receptors for Light and a host of proteins present in spinal fluid for Schutzer. And they differ in design – experimental study of fatigue for Light and descriptive study for Schutzer. Surprisingly, none of the proteins listed for CFS would be products of the gene expression differences reported by Light and colleagues. However, that is not where the story should end.

Several of the “pathways” or clusters of proteins that Schutzer and colleagues reported as uniquely represented in CFS are potentially the same pathways involved in the gene expression differences reported by Light and colleagues. For example, the adrenergic pathways studied by Schutzer have relevance for adrenergic gene expression results reported by the Lights’ team. Finally, both studies have provided critical data necessary for future research. The data from Schutzer and colleagues provides a plethora of potential protein targets for future validation studies and the identified pathways should help guide future hypothesis-driven research. The data from Light and colleagues could lead to both objective diagnostic markers and potential targets for treatment interventions.

As for the second question about the sample size necessary to validate the Light data, that depends on the design of the study and the magnitude of the expected results. Bigger isn’t always better and in research we strive to strike a balance between having as many participants as necessary to power our studies without subjecting more participants than necessary to the sometimes demanding experimental protocols. For example, you may need only a relatively small number of participants if you expect that patients and controls will differ greatly on the variables you are interested in. On the other hand, you may need a large sample size if you want your results to generalize to a certain population of interest and the magnitude of the difference you are interested in tends to be moderate. Independent replication is a critical element of the scientific method and is necessary to validate important findings such as those by Light and colleagues. The numbers needed are a function of the research question. 

References:

Light AR, Bateman L, Jo D, Hughen RW, VanHaitsma TA, White AL, Light KC. Gene expression alterations at baseline following moderate exercise in patients with chronic fatigue syndrome and fibromyalgia syndrome. Journal of Internal Medicine.2011 May 26. doi: 10.1111/j.1365-2796.2011.02405.x.

McCleary KK. Exercise challenge reveals potential CFS biomarkers. Research1st (the CFIDS Association of America). 2011 June 2. http://www.research1st.com/2011/06/02/exercise-challenge-reveals-potential-cfs-biomarkers/

Light AR. Shedding light on biomarkers. Research1st (the CFIDS Association of America). 2 August 2011. http://www.research1st.com/2011/08/02/shedding-light-on-biomarkers/

Schutzer SE, Angel TE, Liu T, Schepmoes AA, Clauss TR, Adkins NJ, Camp DG, Holland BK, Bergquist J, Coyle PK, Smith RD, Fallon BA, Natelson BH. Distinct cerebrospinal fluid proteomes differentiate post-treatment Lyme disease from chronic fatigue syndrome. PLoS ONE 6(2): e17287.  2011 February 23. doi:10.1371/journal.pone.0017287 http://www.plosone.org/article/info%3Adoi%2F10.1371%2Fjournal.pone.0017287

Vernon SD and McCleary KK. The promise of the proteome. CFIDSLink (the CFIDS Association of America). 2011 February 23. http://www.cfids.org/research/proteome-analysis.pdf

Dane B. Cook, Ph.D., is an assistant professor of kinesiology in the department of kinesiology at the University of Wisconsin in Madison. Dr. Cook completed his post-doctoral studies at UMDNJ with Dr. Benjamin Natelson’s team, where he was first introduced to CFS and participated in CFS research projects. Dr. Cook is a secretarial appointee to the Department of Health and Human Services CFS Advisory Committee and he has received several other awards, including a New Investigator Award from the American College of Sports Medicine. He is a grantee of the CFIDS Association of America under its Research Institute Without Walls; he will lead the study, “Post-exertion Malaise in CFS/ME: Brain, inflammation and behavioral interactions.” Read more about it at http://bit.ly/2012-cook

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