Our results indicate the existence of a basolateral short-chain fatty acid/HCO₃⁻ exchanger, with monocarboxylate transporter 1 as a primary candidate for acetate transfer.

Butyrate modulates the differentiation, proliferation and gene expression profiles of various cell types. Ruminal epithelium is exposed to a high intraluminal concentration and inflow of n-butyrate. We aimed to investigate the influence of n-butyrate on the mRNA expression of proteins involved in the transmembranal transfer of n-butyrate metabolites and short-chain fatty acids in ruminal epithelium. N-butyrate-induced changes were compared with the effects of hypoxia because metabolite accumulation after O2 depletion is at least partly comparable to the accumulation of metabolites after n-butyrate exposure. Furthermore, in various tissues, O2 depletion modulates the expression of transport proteins that are also involved in the extrusion of metabolites derived from n-butyrate breakdown in ruminal epithelium. Sheep ruminal epithelia mounted in Ussing chambers were exposed to 50 mM n-butyrate or incubated under hypoxic conditions for 6 h. Electrophysiological measurements showed hypoxia-induced damage in the epithelia. The mRNA expression levels of monocarboxylate transporters (MCT) 1 and 4, anion exchanger (AE) 2, downregulated in adenoma (DRA), putative anion transporter (PAT) 1 and glucose transporter (GLUT) 1 were assessed by RT-qPCR. We also examined the mRNA expression of nuclear factor (NF) κB, cyclooxygenase (COX) 2, hypoxia-inducible factor (HIF) 1α and acyl-CoA oxidase (ACO) to elucidate the possible signalling pathways involved in the modulation of gene expression. The mRNA expression levels of MCT 1, MCT 4, GLUT 1, HIF 1α and COX 2 were upregulated after both n-butyrate exposure and hypoxia. ACO and PAT 1 were upregulated only after n-butyrate incubation. Upregulation of both MCT isoforms and NFκB after n-butyrate incubation could be detected on protein level as well. Our study suggests key roles for MCT 1 and 4 in the adaptation to an increased intracellular load of metabolites, whereas an involvement of PAT 1 in the transport of n-butyrate also seems possible.

In the intact rumen epithelium, isoforms 1 and 4 of the monocarboxylate transporter (MCT1 and MCT4) are thought to play key roles in mediating transcellular and intracellular permeation of short-chain fatty acids and their metabolites and in maintaining intracellular pH. We examined whether both MCT1 and MCT4 are expressed at mRNA and protein levels in ovine ruminal epithelial cells (REC) maintained in primary culture and whether they are regulated by peroxisome proliferator-activated receptor-α (PPARα). Because both transporters have been characterized to function coupled to protons, the influence of PPARα on the recovery of intracellular pH after l-lactate exposure was evaluated by spectrofluorometry. MCT1 and MCT4 were detected using immunocytochemistry both at the cell margins and intracellularly in cultured REC. To test regulation by PPARα, cells were exposed to WY 14.643, a selective ligand of PPARα, for 48 h. The subsequent qPCR analysis resulted in a dose-dependent upregulation of MCT1 and PPARα target genes, whereas response of MCT4 was not uniform. Protein expression of MCT1 and MCT4 quantified by Western blot analysis was not altered by WY 14.643 treatment. l-Lactate-dependent proton export was blocked almost completely by pHMB, a specific inhibitor of MCT1 and MCT4. However, l-lactate-dependent, pHMB-inhibited proton export in WY 14.643-treated cells was not significantly altered compared with cells not treated with WY 14.643. These data suggest that PPARα is particularly regulating MCT1 but not MCT4 expression. Extent of lactate-coupled proton export indicates that MCT1 is already working on a high level even under unstimulated conditions.

Conjugated linoleic acids (CLA), particularly cis-9,trans-11 (c9t11) and trans-10,cis-12 (t10c12), are used as feed additives to adapt to constantly increasing demands on the performance of lactating cows. Under these feeding conditions, the rumen wall, and the rumen epithelial cells (REC) in particular, are directly exposed to high amounts of CLA. This study determined the effect of CLA on the fatty acid (FA) metabolism of REC and expression of genes known to be modulated by FA. Cultured REC were incubated with c9t11, t10c12, and the structurally similar FA linoleic acid (LA), oleic acid (OA), and trans-vaccenic acid (TVA) for 48 h at a concentration of 100 µM. Cellular FA levels were determined by gas chromatography. Messenger RNA expression levels of stearoyl-CoA desaturase (SCD) and monocarboxylate transporter (MCT) 1 and 4 were quantified by reverse transcription-quantitative PCR. Fatty acid evaluation revealed significant effects of CLA, LA, OA, and TVA on the amount of FA metabolites of β-oxidation and elongation and of metabolites related to desaturation by SCD. The observed changes in FA content point (among others) to the ability of REC to synthesize c9t11 from TVA endogenously. The mRNA expression levels of SCD identified a decrease after CLA, LA, OA, or TVA treatment. In line with the changes in mRNA expression, we found reduced amounts of C16:1n-7 cis-9 and C18:1n-9 cis-9, the main products of SCD. The expression of MCT1 mRNA increased after c9t11 and t10c12 treatment, and CLA c9t11 induced an upregulation of MCT4. Application of peroxisome proliferator-activated receptor (PPAR) α antagonist suggested that activation of PPARα is involved in the changes of MCT1, MCT4, and SCD mRNA expression induced by c9t11. Participation of PPARγ in the changes of MCT1 and SCD mRNA expression was shown by the application of the respective antagonist. The study demonstrates that exposure to CLA affects both FA metabolism and regulatory pathways within REC.