B Cell Maturation
Longitudinal StudyLongitudinal Metabolic and Genomic Study
Aim: Monitor how metabolic pathways change in ME/CFS patients over a short-term stress event and over the long-term maintenance of the disorder.
All previous metabolomics research on ME/CFS has viewed the metabolites in a single urine, blood and fecal sample from a cohort of patients with ME/CFS and non-ME/CFS controls, many significant differences were observed between the two groups and we have highlighted biological pathways that we hypothesise to be significant to the disorder. We previously have observed energy metabolism and oxidative stress issues are apparent in ME/CFS and that the energy metabolism in particularare associated with gut microbiota and metabolite changes. The next step is to determine how these pathways interact and whether they are causal or symptomatic of ME/CFS. To do this we plan to look at the metabolite changes over multiple sampling points in time from each individual while also viewing the genes that are being expressed over those sample points.
We aim to take several samples in short succession during a flu event and a physical exercise stress event. We hope to observe how metabolism changes in patients with ME/CFS when dealing with stress events. We hypothesise that the change in metabolism in response to stress and the recovery from a stress event may be a key descriptor of ME/CFS. We also aim to take samples every month over a 6-month period to assess long-term changes in metabolism in ME/CFS.
With this work we hope to uncover more details about the pathogenesis of ME/CFS. We aim to relate metabolic pathways to the symptoms experienced in each individual and consequently discovering common pathways in the whole ME/CFS cohort that differ from those observed in the non-ME/CFS control group.
As aluded to earlier, our previous studies were biomarker-driven and these led to the observation of pathways that may be significant to the disorder but weren’t 100% descriptive of the disorder based on the single-point analysis. Changes observed from single-point analysis are difficult to be defined as causal or symptomatic, by observing the data change over time we aim to observe trigger points and the development of metabolic changes that originate at the genetic level.
A secondary aim is a proof-of-principle for the development of this workflow to effectively monitor the biochemical pathways in ME/CFS patients and how they may react to different stress-situations and also how they may improve. By monitoring patient symptoms we will be able to observe time-points of improved symptoms and relate that to the biochemistry changes. Developing this type of technology could be crucial to creating tailored treatment options and monitoring the effectiveness of treatments for patients with ME/CFS.
The significance of this study is the potential to find a causal link to the energy metabolism, amino acid metabolism, oxidative stress and gut dysbiosis issues that we have observed in ME/CFS patients. We already have novel work completed in this field with intriguing resultsand we are looking to expand upon them to reach their ultimate conclusion. This study will also provide a framework for the future of organizing and monitoring personalised medicine in treating ME/CFS as we monitor the improvements in ME/CFS and its relationship to the individual’s underlying biochemistry.
B Cell MaturationMetabolic Analysis of B Cell Maturation
Collaboration project with Fane Mensah and Professor Jo Cambridge at University College London
Aim: Monitor the metabolic requirements of B cell maturation to observe if altered metabolic environments impact B cell maturation and, conversely, if B cell maturation alters the metabolic environment.
Previous research has found consistent evidence of abnormalities in the immune system, energy metabolism and oxidative stress in ME/CFS patients. Promising improvement after B-cell depletion therapy with Rituximab highlighted the possible involvement of the immune system and (in this case B cells) in ME/CFS.
Fane Mensah and Jo Cambridge have found abnormalities in the maturation of B cells, observing phenotypic markers on B cell subsets and anomalies of mitochondrial mass. We now want to find out the metabolic requirements for B cells and to see if there are differences between the metabolism of ME/CFS and non-ME/CFS B cells. We also want to know if the B cells impact the metabolic environment and conversely if the metabolic environment impacts the B cells.
This study is looking at two key areas of dysfunction in ME/CFS: energy metabolism and the immune system. Consistent findings have been observed in both and links have been made between the two but there is yet a study that has combined them to this level of detail. The relevance of this study to ME/CFS is that it attempts to determine the origin of mitochondrial dysfunction in B cells and how this may be the determinant for immune dysfunction and more broadly the disorder itself.
We propose that the systemic features found in ME/CFS are due to chronic changes affecting the homeostasis of immune and somatic cells. Metabolic thresholds induced following stressors such as viral infections may be altered, resulting in disturbed cytokine profiles and interactions between these cells. By investigating the mRNA and metabolites produced during B cell maturation and proliferation in vitro we will determine the energy requirement (and possible dysfunction) of different states of cellular metabolism that may be significant to ME/CFS. By performing a genome analysis and self-reporting of patient symptoms prior to and at the point of sample collection we will be able to stratify patients and associate anomalies in cellular metabolism to the broad and specific clinical features associated with ME/CFS.
*Image: Blausen.com staff (2014). "Medical gallery of Blausen Medical 2014". WikiJournal of Medicine 1 (2). DOI: 10.15347/wjm/2014.010.
MitochondriaMitochondrial Respiratory Function and Cellular Metabolism in Lymphoblasts
Collaboration project with Daniel Missailidisand Professor Paul Fisher at LaTrobe University
Aim: Evaluate mitochondrial and cellular energy pathways to resolve the foundation of the energy anomalies that have been observed from metabolic studies on ME/CFS patients.
Mitochondria are structures present within the cells of our body and most prominently produce ATP via aerobic respiration – this is akey component of cellular energy supply. Mitochondrial energy production relies upon oxygen and has several key pathways that lead to energy production. We can measure oxygen consumption by cells in real time, which enables the input of enzyme inhibitors of mitochondrial processes to be added. The end-product of such an assay is to see the maximum energy production from mitochondria as it loses important pathways step-by-step. This allows us to pinpoint the areas in the mitochondria that differ between ME/CFS and non-ME/CFS groups.
We plan to run these experiments in conjunction with metabolite analysis of the blood and cells as well. This will enable us to observe the metabolite changes in energy pathways external to the mitochondria. We will also measure important regulatory molecules of ATP production in cells,AMPK (AMP-activated protein kinase) and/or TORC1 (TOR Complex I).
This work will be conducted upon immortalised white blood cells (referred to as lymphoblasts) that are isolated from ME/CFS patient and control blood. We hope that the results will provide important information on the pathogenesis of ME/CFS while testing the hypothesis that the disorder involves altered energy pathways, elevated levels of cellular oxidative stress and dysregulation of cellular stress-sensing signalling pathways.
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