Short-Chain Fatty Acids in Lypopolysaccharide-Induced Inflammation and Executive Function

INFLEX is a single-center, randomized, triple-blind, placebo-controlled, parallel-group interventional trial conducted at UZ Leuven / KU Leuven. Its primary aim is to test whether a single acute dose of colon-delivered SCFAs, relative to placebo, attenuates the systemic inflammatory response induced by experimental endotoxemia in healthy adults. Secondary aims are to test whether SCFAs attenuate LPS-induced sickness behaviour and mitigate endotoxemia-induced changes in core executive functioning, and to characterize associations between systemic SCFA uptake, the inflammatory response, and these outcomes.

Eligible participants are randomized to receive SCFA colon-delivery capsule or placebo capsules, taken on the morning of the test day with a standardized low-fiber breakfast. One hour after capsule intake, participants receive a single intravenous bolus of purified Escherichia coli lipopolysaccharide (0.4 ng/kg body weight), which serves as a fixed inflammatory challenge applied to all participants. The timing is designed so that peak circulating SCFA concentrations (approximately 3-4 hours after colonic delivery) coincide with the peak of LPS-induced cytokines and sickness behaviour (approximately 2-3 hours post-injection).

Participants are monitored for a minimum of 6 hours after the LPS challenge. Vital signs, sickness symptoms, and blood and saliva samples are collected hourly to quantify inflammatory markers (including IL-6 and hs-CRP), serum SCFA, cortisol, and ACTH. Core executive-function tasks are administered before and after the LPS challenge and at a 24-hour follow-up visit. A fecal sample collected at home prior to the test day is used for gut microbiota profiling. To standardize endogenous SCFA production, participants follow a low-fiber diet for three days before the test day and avoid caffeine, alcohol, and strenuous exercise beforehand. The night prior to the test day, participants are instructed to consume a standard dinner that is provided to them.

The study comprises three visits: a screening visit (~1 hour), the laboratory test day (~8 hours), and a follow-up visit (~30 minutes) 24 hours after the LPS challenge.

Inclusion criteria

• Voluntary written informed consent obtained prior to any study procedure
• Healthy, with no gastrointestinal or psychological complaints
• Age 18-45 years
• BMI 18.5-27 kg/m2
• Proficiency in English and/or Dutch

Exclusion criteria

• History of previous or current neurological disorder (e.g., epilepsy, multiple sclerosis, migraine with aura)
• History of previous or current psychiatric disorder (e.g., depression, anxiety disorders, bipolar disorder, schizophrenia)
• History of previous or current gastrointestinal disorder (e.g., Crohn's disease, ulcerative colitis, irritable bowel syndrome)
• History of previous or current endocrine disorder (e.g., diabetes, thyroid disorders, polycystic ovary syndrome)
• History of immune system disorder, including inflammatory, autoimmune, or severe allergic conditions (e.g., rheumatoid arthritis, lupus, celiac disease, or severe allergies such as anaphylaxis)
• History of neuropsychiatric disorder (e.g., ADHD, autism spectrum disorder, learning disorder, Tourette's syndrome)
• History of major head trauma (e.g., concussion with loss of consciousness or long-term symptoms)
• History of severe infection (e.g., sepsis, pneumonia requiring hospitalization, meningitis, endocarditis, or other systemic infection requiring hospitalization, intravenous antibiotics, or intensive care)
• One or more diagnoses based on the Mini International Neuropsychiatric Interview (MINI)
• One or more diagnoses based on ROME-IV criteria for gastrointestinal disorders
• Any disorder that, in the investigator's opinion, might jeopardise the participant's safety or compliance with the protocol
• Any prior or concomitant treatment that might jeopardise the participant's safety or compromise the integrity of the study
• Participation in an interventional trial with an investigational medicinal product (IMP) or device
• Current or recent (past month) prescribed medication use, with particular attention to cardiovascular drugs, steroids, NSAIDs, and centrally-acting drugs (excluding isolated over-the-counter use such as ibuprofen or paracetamol, and hormonal contraceptives in women)
• Use of psychotropic medication within the past year (e.g., antidepressants, anxiolytics, antipsychotics)
• Use of antibiotics within three months preceding the study
• Use of pre- or probiotics within one month preceding the study
• Current or recent (past month) infection (e.g., common cold, influenza, COVID-19)
• Recent (past month) vaccination
• Pregnant or lactating women
• Previous or current substance or alcohol dependence or abuse (>2 units/day or >14 units/week)
• Smoking
• Night-shift work
• Adherence to special diets (e.g., vegan, vegetarian, weight-loss, lactose-free, gluten-free)
• Previous experience with or knowledge of any of the cognitive tasks used in the study (questionnaires excluded)
• C-reactive protein higher than 0.5mg/dL on the day of the injection.
• Colour vision deficiency or colour-blindness