Deciphering the Impact of Exposures From the Gut Microbiome-derived Molecular Complex in Human Health and Disease (ExpoBiome)

Deciphering the Impact of Exposures From the Gut Microbiome-derived Molecular Complex in Human Health and Disease

Deciphering the Impact of Exposures From the Gut Microbiome-derived Molecular Complex in Human Health and Disease

The ExpoBiome project will analyze the impact of fasting on patients with Parkinsons's Disease (PD) or rheumatoid arthritis (RA) on a clinical level as well as the effect of fasting on their immune system and gut microbiota. ExpoBiome will combine metagenomics and other "omics" [meta-transcriptomics, meta-proteomics and (meta-)metabolomics], bioinformatic analyses and biostatistics under a systems biology framework to gain new mechanistic insights into microbiome-immune system interactions in the context of chronic diseases with inflammatory signatures. Besides a one time crossectional study of healthy participants, patients with RA and PD a longitudinal fasting study with two arms (RA and PD) is planned.

The human gut microbiome is a complex ecosystem, which contributes essential functions to human physiology. Changes to the microbiome are associated with several chronic diseases characterised by inflammation, including neurodegenerative and autoimmune diseases. Microbiome-derived effector molecules comprising nucleic acids, (poly)peptides and metabolites are present at high levels in the gut but have so far eluded systematic study. This gap in knowledge is limiting mechanistic understanding of the microbiome's functional impact on chronic diseases such as Parkinson's Disease (PD) and rheumatoid arthritis (RA). Here, for the first time a combination of advanced high-resolution methodologies will be integrated to comprehensively identify the constituents of this molecular complex and their impact on the human immune system. First, a quantitative, integrated multi-omic analysis on microbiome samples collected from healthy individuals and patients with newly diagnosed PD or RA will be performes. Using contextualised prior knowledge (ExpoBiome Map) and machine learning methods, we will identify microbial molecules associated with condition-specific immunophenotypes. Second, the biomarker signature during a model clinical intervention (therapeutic fasting) will be validated and tracked to predict treatment outcomes. Third, microbes and molecules will be screened in personalised HuMiX gut-on-chip models to identify novel anti-inflammatory compounds. By providing mechanistic insights into the molecular basis of human-microbiome interactions, the project will generate essential new knowledge about causal relationships between the gut microbiome and the immune system in health and disease. By facilitating the elucidation of currently unknown microbiome-derived molecules, it will identify new genes, proteins,metabolites and host pathways for the development of future diagnostic and therapeutic applications.

Patients undergo a 5-10 day fasting period with a dietary energy supply 350-400kcal per day with fruit and vegetable juices or, if not feasible, an established fasting-mimicking diet of 600-800 kcal according to Longo et al.

Molecular typing of the gut microbiota using sequencing and high-throughput analysis from stool samples (metagenomics, metatranscriptomics, metaproteomics, metabolomics)

Change from Baseline in the DAS-28-CRP, range from 2.0 to 10.0 while higher values meaning a higher disease activity and below of 2.6 meaning remission

Change from Baseline in the HAQ, range from 0 to 3 while higher values meaning a higher grade of disability

Change from Baseline in the SDAI, range from 0 to 86 with assumed range from 0.1 to 10mg/dL for CRP. Higher values mean a higher disease activity and below of 34 meaning remission.

Change from Baseline in the FFbH-R, range from 0 to 100 % while higher values meaning a higher grade of functional capacity

Movement Disorders Society-sponsored revision of the Unified Parkinson's Disease Rating Scale part I, II, IV (MDS-UPDRS part I, II, IV). Total score range: 0-260 (including the MDS-UPDRS part III), including 4 subcategories. Subcategories are summed to compute the total score. A lower score means a better outcome.

The Parkinson's Disease Sleep Scale 2 (PDSS-2) is designed to assess nocturnal disability in Parkinson's disease. The PDSS-2 is a 15 question analog scale that ranks answers from 0 - 4, with 4 being worse. (Question 1 is an exception, where 4 is better and 0 is worse). In addition to an overall assessment of sleep disability three aspects of sleep problems can be obtained; disturbed sleep (total of questions 1-3, 8 and 14), PD-specific nocturnal motor symptoms (total of questions 4-6, 12 and 13), and PD-specific nocturnal symptoms (Total of questions 7, 9-11 and 15).

The 39 question Parkinson's Disease Questionnaire (PDQ-39) is a patient-reported rating scale for quality of life in Parkinson's disease. Respondents affirm if they have experienced problems due to their disease using a five point scale from never (0 points) to always (4 points, or worse) in doing common activities. The PDQ-39 is comprised of 8 domains: mobility, emotion, activities of daily living, cognition, stigma, social support, communication, bodily discomfort. Total possible range of scores = 0 - 156, with higher scores representing worse severity.

The non-motor symptoms (NMS) questionnaire can be given to people affected by Parkinson's in order to aid health and social care professionals to assess their non-motor symptoms.The non-motor symptoms questionnaire is a 30-point, patient-based questionnaire used to determine the non-motor symptoms experienced by the patient during the past month. The points should be totalled to give a score out of 30. A score of under 10 is mild, 10-20 moderate and over 20, severe.

The Non-Motor Symptoms Scale (NMSS) measures the frequency and severity of a range of non-motor symptoms in Parkinson's Disease, through 30 questions grouped into 9 domains: cardiovascular, sleep/fatigue, mood/cognition, perceptual problems/hallucinations, attention/memory, gastro-intestinal tract, urinary, sexual function, and miscellaneous (pain, taste/smell, weight change, excessive sweating). Severity is rated on a scale from 0 (none) to 3 (severe). Frequency is rated on a scale from 1 (rarely) to 4 (very frequent). Item scores are calculated as the product of severity and frequency; the total score is obtained by summing the item scores. The NMSS total score ranges from 0 to 360 with a lower score indicating fewer symptoms. A negative change from baseline indicates improvement in symptoms (reduced score).

Change from Baseline in the CPSS, range from 0 to 4 in each item. Scores are obtained by reversing responses (e.g., 0 = 4, 1 = 3, 2 = 2, 3 = 1 & 4 = 0) to the positively stated items and then summing across all scale items, higher values meaning a higher grade of perceived stress.

Change from Baseline in the WHO-5, range from 0 to 100 %, higher values meaning a higher grade of well-being

Change from Baseline in Emotional Distress will be measured using the German Version of the Profile of Mood States (ASTS) short version (19 items, 7-point Likert scale; 0=not at all, 6=extremely). Lower scores indicate more stable mood profiles.

Age, gender, education level, household income, employment status, marital status, language spoken, complete family history, current and previous illness and co-morbidities, and current medications

Physical inactivity, coffee, health promoting activities via Likert Scales, range from 0 to 5 while higher values meaning a higher grade of agreement

Modified FFQ recording dietary behaviour such as mealtimes, frequency of food intake, food preferences, fasting experiences

Determination of cytometric parameters that indicate changes in cell activation or quantitative changes in the absolute and/or relative size of subpopulations (e.g. classical/intermediate/non-classical monocytes, naïve and memory T-cells, B-cell differentiation to plasmablasts/-cells) Gene expression analysis of immune cells with Affymetrix whole genome microarrays and RNAseq to search for transcriptional patterns and markers that help to identify relevant immune cell (sub-)populations, which are not yet included in the cytometric phenotyping screen

Inclusion Criteria: One of the following diagnoses: rheumatoid arthritis (first diagnosis >6 weeks ago and <8 years), parkinson's disease OR healthy volunteer Control ("healthy") individuals must be without any evidence of active, known or treated RA, without any evidence of active, known or treated central nervous system disease, and without a known family history of idiopathic PD Arthritis in at least one joint Control individuals should match the RA or PD individuals as closely as possible, especially their age, sex, and education Present written declaration of consent Consent to specimen collection and specimen use Ability to understand the patient information and willingness to sign the consent form Exclusion Criteria: gout or proven bacterial arthritis Psychiatric illness that limits understanding of the examination protocol (unable to consent) BMI < 18.5 Pre-existing/current eating disorders (bulimia nervosa, anorexia nervosa) within the past 5 years. Severe internal diseases (e.g. renal insufficiency with creatinine > 2mg/dl) Participation in another study Existing vegan diet or fasting within the past 6 months Pregnancy or breastfeeding Chronic inflammatory bowel disease Use of antibiotics within the past 12 months Presence of anemia