Location: Immunity and Disease Prevention Research
Project Number: 2032-51530-026-04-T
Project Type: Trust Fund Cooperative Agreement
Start Date: Jan 1, 2019
End Date: Sep 30, 2021
Objective 1: Using dietary data and fecal samples from the Western Human Nutrition Center (WHNRC) Phenotyping Study, determine the association of total dairy intake or dairy intake types (fluid milk, cheese, yogurt) with markers of gastrointestinal inflammation. Objective 2: Using dietary data and fecal samples from the WHNRC’s Phenotyping Study, determine the association of total dairy intake or dairy intake types (fluid milk, cheese, yogurt) with markers of gastrointestinal protection.
Study Design The Phenotyping Study (ClinicalTrials.gov: NCT02367287) is an observational, crossover trial that began in 2015 and will complete enrollment by March 2019 with a target of 396 subjects. Dietary data (Food frequency questionnaire (FFQ) and 3-4 24 hour recalls per subject) and stool samples from this cohort will be used to complete the objectives in the current proposal. Participants: Healthy men and women are being recruited to fill nine recruitment bins, within sex, to balance body mass index (BMI) and age, using three BMI categories (<25, 25–29, 30–40 kg/m2) within each age category (18–33, 34–49, and 50–65 years). Participants whose dietary intake measures (FFQ and 24-hour recalls) pass quality control and who provide a stool sample will be included in the proposed study. We expect 300 participants will meet these inclusion criteria. For the RNA markers of mucin production, only subjects who have provided an RNA-preserved stool sample (expect n=100) will be included. Outcome measures: Fecal calprotectin, myeloperoxidase, neopterin, and sIgA will be measured in duplicate by enzyme-linked immunosorbent assay (ELISA) (Immunodiagnostik). Plasma lipopolysaccharide (LPS)-binding protein will also be measured by ELISA. There are no ELISAs that can be used to measure fecal mucin in humans. Therefore, we will measure messenger RNA (mRNA) markers of mucin production in the subset of study participants who have provided RNA-preserved stool. RNA will be extracted from stool and enriched for polyA transcripts; mucin transcripts will then be quantitated using TaqMan gene expression assays. Our preliminary data from human fecal metatranscriptomics (6 subjects, 4 samples per subject) shows that mRNA for mucin-2 has low variance within subject and high variance between subjects and is therefore likely to be a sensitive and novel marker for phenotyping of gastrointestinal protection.