A Taxonomic Approach to Understanding Diet-Induced Changes in Mouse Fecal and Ileal Immunoglobulin A (IgA) Concentrations Público
Avila, Walter (Spring 2023)
Abstract
The gut microbiome influences immunity and mucosal barrier maintenance and processes in the small intestine. Changes in diet alter the gut microbiota composition, impacting the gut immune system and its constituents. Immunoglobulin A (IgA), an antibody secreted by plasma cells in the intestine that helps defend against harmful pathogens, is also impacted by dietary change. Our preliminary research has shown that mice weaned to a low-fermentable-fiber only (LFF) diet have significantly lower concentrations of ileal and fecal IgA than mice born and maintained on a standard chow diet (SCD). However, the mechanisms underlying this change remain largely unclear. One explanation could be that B cells in the guts of LFF mice produce less IgA in response to the LFF diet. It is also possible that the LFF diet significantly alters the gut microbiome composition, decreasing the repertoire of clonal plasma cells.
To test these hypotheses, we incubated LFF and SCD mouse ileal and fecal contents with IgA-targeting antibodies. These were bound to IgA attached to ileal bacteria, identifying them with a fluorescent signal and allowing them to be sorted. Sorted groups will undergo 16s rRNA sequencing to identify the attached bacteria. If no significant differences are found in the bacteria bound by IgA in LFF or SCD ileum and feces contents, this will indicate possible overall IgA reduction because of dietary change. Conversely, if differences are found, this will imply that the B cell repertoire is affected by dietary modification-induced microbiota changes.
To uncover which bacteria orders may impact IgA production, we produced predictive LASSO models that showed that the following bacterial orders had the highest explanatory power in predicting Lamina propria (LP) IgA plasma and B cell frequencies out of total lymphocytes in SCD and LFF mice: CyanobacteriaMLE1-12, EW055, Clostridiales, Enterobacteriales, and Xanthomonadales. Our modeling results indicate that these bacteria orders significantly explain IgA-producing immune cell frequencies and their roles will be investigated further when analyzing the 16s sequencing data from our experiment. Ultimately, this experiment will provide insight into how the absence of dietary fiber affects IgA expression and influences its role in defending against gut pathogens.
Table of Contents
TITLE PAGE 1
SUMMARY 2
INTRODUCTION 3
RESEARCH QUESTION AND HYPOTHESES 5
BACTERIAL SORT-EXPERIMENTAL DESIGN 7
BACTERIAL SORT-PROTOCOL STANDARDIZATION (PS) 8
STANDARDIZED FINAL BACTERIAL SORT EXPERIMENTS 46
CELL-SORTING RESULTS FROM BACTERIAL-SORT EXPERIMENTS 47
A PREDICTIVE MODEL FOR IGA PLASMA CELL AND B CELL FREQUENCIES GIVEN BACTERIA ORDER RELATIVE PROPORTIONS 56
PREDICTIVE MODEL CONSTRUCTION 58
PREDICTIVE MODEL RESULTS 60
PREDICTIVE MODEL SUMMARY 62
DISCUSSION AND FUTURE DIRECTIONS 63
MATERIALS AND METHODS SUMMARY 65
AUTHOR CONTRIBUTIONS 71
DECLARATION OF INTERESTS 72
DATA AVAILABILITY 72
REFERENCES 72
SUPPLEMENTARY INFORMATION 75
SUPPLEMENTARY FIGURES 76
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