Abstract
Background/Objectives: Dietary compounds simultaneously act on host tissue and gut microbiota, complicating efforts to distinguish direct host–receptor mechanisms from microbiome-mediated effects. Using a common food ingredient as proof-of-principle, this study aims to disentangle capsaicin host receptor signalling from capsaicin–microbiome interactions to determine the role of the gut microbiome in capsaicin-mediated metabolic protection. Methods: We employed a high-fat diet (HFD)-induced obesity model with or without capsaicin supplementation in TRPV1 knockout mice to determine the role of the host receptor. We also depleted the gut microbiota with antibiotics to determine the role of the microbiome. Host metabolic phenotypes were monitored, and microbiome 16S rRNA sequencing and metabolomics were analysed and extended with functional prediction and machine learning algorithms, followed by functional validation in vivo. Results: Capsaicin supplementation significantly protected against HFD-induced obesity, even capsaicin receptor knockout hosts. Crucially, depletion of the microbiota abolished this metabolic protection, demonstrating the microbiome’s obligatory role. While capsaicin supplementation did not alter hosts’ cumulative food intake, dietary preferences, or energy expenditure, it induced significant shifts in gut microbial diversity and function and triggered extensive faecal metabolomic remodelling which indicate altered nutrient processing including dampened lipid absorption. Consistent with this finding, in vivo functional validation also reveals attenuated circulating lipid response following oral fat administration, and this effect of capsaicin is abrogated without the microbiome. Conclusions: Although capsaicin is well-recognised as a modulator of host appetite and metabolism via canonical host receptor signalling, our strategic use of a host knockout has enabled us to unmask a nuanced microbiome-modulatory role of this chilli pepper ingredient: capsaicin tweaks the microbiome to potentially dial down host fat absorption and protect against high-fat diet-induced obesity. We also highlight a generalisable host knockout experimental framework for distinguishing microbiome-mediated pathways versus direct host receptor mechanisms.