Characterization of butyrate transport across the luminal membranes of equine large intestine.

The researchers isolated membranes from horse colon cells to understand how butyrate, a short-chain fatty acid produced by gut bacteria during the fermentation of grass, is transported into the cells. They have identified that the protein monocarboxylate transporter 1 (MCT1) predominantly facilitates butyrate transport across the cellular membrane.

Background

• The study was motivated by the importance of the dietary intake of horses, primarily grass, which is fermented by the microbes in horse’s large intestine to produce short-chain fatty acids. These fatty acids, particularly butyrate, not only provide a major energy source for horses, but also participate in many crucial physiological processes.
• In this context, understanding the mechanism and transport of butyrate into the cells across the luminal membrane (the inner layer of the intestine exposed to the gut contents) is of significant interest.

Experimental Approach

• They began by isolating luminal membrane vesicles from horse colonic cells. These vesicles are essentially ‘bladders’ of cell membrane that allow testing of transport processes in isolation.
• The researchers then analysed these vesicles for different proteins including intestinal luminal membrane markers (Cysteine-sensitive alkaline phosphatase and villin), a glucose transporter (GLUT2) and a pump protein (Na(+)-K(+)-ATPase). These investigations confirmed that the vesicles represent the luminal membrane.
• Furthermore, the researchers used immunohistochemistry to show that the MCT1 protein is present on the luminal membrane of the cells in the horse colon (colonocytes).

Results and Interpretation

• The research team showed that butyrate transport is driven by a pH gradient and is not dependent on sodium (Na+). It is also time and concentration dependent, with specific kinetics that are indicative of a particular transport mechanism.
• Butyrate transport into the vesicles was significantly blocked by p-chloromercuribenzoate, phloretin and α-cyano-4-hydroxycinnamic acid – all known inhibitors of MCT1. This suggests that MCT1 facilitates the transport of butyrate.
• Transport was also inhibited by other short-chain fatty acids and monocarboxylates (compounds with one carboxylate group), further supporting the involvement of MCT1, known to transport these substances.
• All these findings strongly suggest that butyrate is mainly transported across the equine colonic luminal membrane via the MCT1 protein.