Transport of ketone bodies and lactate in the sheep ruminal epithelium by monocarboxylate transporter 1

Abstract

Due to intensive intracellular metabolism of short-chain fatty acids, ruminal epithelial cells generate large amounts of D-beta-hydroxybutyric acid, acetoacetic acid, and lactic acid. These acids have to be extruded from the cytosol to avoid disturbances of intracellular pH (pH(i)). To evaluate acid extrusion, pH(i) was studied in cultured ruminal epithelial cells of sheep using the pH-sensitive fluorescent dye 2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein. Extracellular addition of D-beta-hydroxybutyrate, acetoacetate, or lactate (20 mM) resulted in intracellular acidification. Vice versa, removing extracellular D-beta-hydroxybutyrate, acetoacetate, or lactate after preincubation with the respective monocarboxylate induced an increase of pH(i). Initial rate of pH(i) decrease as well as of pH(i) recovery was strongly inhibited by pCMBS (400 microM) and phloretin (20 microM). Both cultured cells and intact ruminal epithelium were tested for the possible presence of proton-linked monocarboxylate transporter (MCT) on both the mRNA and protein levels. With the use of RT-PCR, mRNA encoding for MCT1 isoform was demonstrated in cultured ruminal epithelial cells and the ruminal epithelium. Immunostaining with MCT1 antibodies intensively labeled cultured ruminal epithelial cells and cells located in the stratum basale of the ruminal epithelium. In conclusion, our data indicate that MCT1 is expressed in the stratum basale of the ruminal epithelium and may function as a main mechanism for removing ketone bodies and lactate together with H+ from the cytosol into the blood.

Gene expression in the digestive tissues of ruminants and their relationships with feeding and digestive processes

“…Maintenance of rumen intracellular pH is accomplished partly through exchange proteins, such as Na 1 /H 1 exchange (NHE) proteins and MCT1 that regulate cellular H 1 efflux (Gä bel and Aschenbach, 2006) and transport of SCFA metabolites into the blood (Mü ller et al, 2002), respectively. The NHE proteins, in concert with Na 1 /K 1 -ATPases (Albrecht et al, 2008), counteract the increase in H 1 concentration in the cytosol during transport of SCFA into the rumen epithelium (Mü ller et al, 2000).…”

“…At the protein level, immunohistochemistry (IHC) studies indicated localized expression of the MCT1 protein almost exclusively in the stratum basale of the stratified epithelial layers of the rumen (Mü ller et al, 2002;Graham et al, 2007) and diffuse, weak staining for MCT2 in the three epithelial layers beneath the stratum corneum (Graham et al, 2007). Using primary cultures of ovine ruminal epithelium treated with b-hydroxybutyrate, acetoacetate and lactate with and without competitive blockers of MCT1, Mü ller et al (2002) demonstrated MCT1-mediated uptake of ketone bodies and lactate. Their uptake was associated with a drop in intracellular pH.…”

“…In bovine rumen, only MCT1 and MCT2 mRNA transcripts were detected. Expression of MCT1 mRNA in ovine rumen epithelium was demonstrated also by Mü ller et al (2002). At the protein level, immunohistochemistry (IHC) studies indicated localized expression of the MCT1 protein almost exclusively in the stratum basale of the stratified epithelial layers of the rumen (Mü ller et al, 2002;Graham et al, 2007) and diffuse, weak staining for MCT2 in the three epithelial layers beneath the stratum corneum (Graham et al, 2007).…”

“…Acetate, propionate and butyrate are the principal SCFA produced by rumen microbes during carbohydrate fermentation and subsequently serve as the principal energy source for ruminants (Bergman, 1990). However, their uptake by epithelial cells lining the alimentary canal is partially dependent upon carriermediated transporters (Mü ller et al, 2002;Graham et al, 2007) in addition to passive diffusion across the apical membrane of epithelial cells (Sehested et al, 1999). The existence of multiple monocarboxylate transporter ( MCT) isoforms has been demonstrated in various mammalian cell types (Halestrap and Price, 1999), and mRNA expression of seven MCT isoforms (MCT1 through MCT7) was evaluated by Graham et al (2007) in bovine rumen epithelium by RT-PCR.…”

Transport of cations and anions across forestomach epithelia: conclusions from in vitro studies

“…The majority of these SCFA (which consist mainly of acetate (,60%), propionate (,20%) and butyrate (,10%) are absorbed across the forestomachs and cover a substantial part of the animals energy requirements (Bergman, 1990). The major fraction of SCFA crosses the forestomach epithelia unmetabolized (Kiddle et al, 1951;Gä bel et al, 2002), although SCFA, in particular butyrate, are also used by the epithelial cells themselves for cell metabolism, which could support active transport of other electrolytes (Weigand et al, 1975;Sehested et al, 1999;Kristensen et al, 2000;Gä bel et al, 2002;Gä bel and Aschenbach, 2006). Apical uptake mechanisms for SCFA Previous studies and reviews have focussed on two possible mechanisms of SCFA uptake across the apical membrane: diffusion of the protonated, undissociated form through the lipid bilayer and exchange of the dissociated SCFA 2 anion for bicarbonate via a transport protein that exchanges SCFA 2 for HCO 3 2 anions under physiological conditions (Figure 4, Gä bel et al, 1991a;Kramer et al, 1996;Gä bel et al, 2002;Aschenbach et al, 2009).…”