Abstract/Summary

Gut barrier dysfunction is associated with several chronic inflammatory, non-infectious diseases which are prevalent in the UK population. Increased gut permeability allows bacteria and bacterial-derived products to pass through to the underlying lamina propria where they can trigger pro-inflammatory responses. Inflammation exacerbates poor gut barrier function and thus, a feedback loop is generated. High protein diets can alter gut microbial population dynamics and subsequent increases in concentrations of protein associated metabolites which have been shown to impair gut barrier function in vitro. Given the sexual dimorphism in microbiota composition, gut barrier function and immune responses, the hypothesis was that sex-by-protein interactions would influence barrier dysfunction. The aim of this PhD was to explore the sex-dependent effects of high protein diets on gut microbiotas, metabolite production and their impacts on gut barrier function and mucosal immunity using both in vitro batch culture systems and pig models. Faecal samples from male (n = 5) and female (n = 5) donors were incubated with protein from a variety of different sources under simulated colonic conditions, while male (n = 10) and female (n = 10) pigs were fed high protein (28% dietary protein; n = 10) or standard protein (18% dietary protein; n = 10) diets for four weeks in a 2x2 trial (n = 5 litter-matched pigs/sex/treatment group). Changes in microbial populations were assessed using fluorescent in situ hybridisation coupled with flow cytometry (FISHflow) or 16S rRNA sequencing techniques, metabolite profiles were determined using gas chromatography / mass spectrometry (GC/MS) and solid-phase microextraction gas chromatography-mass spectrometry (SPME/GC-MS) while barrier function and immune-associated protein expression was quantified using fluorescence immunohistology methods. Significant in vitro effects of protein source and sex were observed on microbiota composition and metabolite production. With additional protein availability, ammonia production by the microbiota from females was significantly higher than that from males (p < 0.01), while p-cresol production was significantly higher in the microbiota from males compared to females (p < 0.001). In pigs, high protein diets drove increased p-cresol production and reduced E-cadherin expression without altering overall microbiota composition. However, females exhibited shifts towards potentially pathogenic genera (Chryseobacterium; p < 0.001, Staphylococcus; p < 0.001) while males had higher Lachnospiraceae_NK4A136 (p = 0.02) and Cloacibacillus (p = 0.016). Notably, female pigs had significant increases in p-cresol (p = 0.049) and reduced ZO-1 expression (p = 0.04), indicating greater vulnerability to diet-induced barrier dysfunction compared to male pigs. These findings suggest sex-specific dietary recommendations may be helpful in reducing the prevalence of barrier dysfunction and related chronic inflammatory diseases.