The effects of FFAR and FFAR
The effects of FFAR1 and FFAR2 activation in different tissues have been reported in several species (Hara et al., 2014). Herein, we report the first evidence for differential expression of FFAR1 and FFAR2 during the peripartal period in the liver of dairy cows that differ in liver metabolism, based on BHB measurement after calving. Our observations regarding the time periods are at least partly in line with Börner et al. (2013). In that study, based on the same animals as used in the study of Schäff et al. (2012), it was observed that cows with more lipid mobilization postpartum showed lower carbohydrate oxidation and higher fat oxidation postpartum but also antepartum. Comparable differences, including the antepartum and postpartum periods, were extended to mRNA levels of genes linked to lipid and carbohydrate metabolic pathways by Schäff et al. (2012), when grouping the cows by liver fat content postpartum. It has been shown, in human HepG2 cells as well as in human primary liver cells, that oleic levosimendan increases the expression of FFAR1 and activates the receptor (Wu et al., 2012). This activation is indirectly linked to improved insulin sensitivity and increased β-oxidation and involves the activation of peroxisome proliferator-activated receptor δ (PPARδ) through the FFAR1–phospholipase C–calcium pathway, as demonstrated in humans and rats. In detail, the increased abundance of PPARδ by increased FFAR1 expression was negatively associated with abundance of phosphatase and tensin homolog (PTEN), which blocks phosphorylation of insulin-dependent serine/threonine protein kinase Akt/PKB. Consequently, the expression of PPARδ by FFAR1 activation improves insulin sensitivity in steatotic cells (Wu et al., 2012). However, it is important to note that differences in liver lipid metabolism between rodents and ruminants exist. For example, in contrast to human or mouse and using glucose as precursor for de novo fatty acid synthesis, the rate of de novo lipogenesis in the liver of ruminants, using mainly acetate as precursor, is low. In addition, the hepatic capacity to esterify fatty acids taken up from the bloodstream is limited (Adewuyi et al., 2005; Bergen and Mersmann, 2005). Less information is available on FFAR1 importance in the bovine. It is known that the FFAR1 agonist GW9508 is functional in the bovine (Manosalva et al., 2015). In mice, the same agonist decreases sterol regulatory element-binding protein 1 (Ou et al., 2014), and overexpression of this transcription factor in bovine hepatocytes in vitro induces triglyceride accumulation (Li et al., 2014). Therefore, because of the functionality of the FFAR1 agonist GW9508 in both species, a comparable effect of FFAR1 on bovine sterol regulatory element-binding protein 1 and thus lipid metabolism cannot be ruled out. We detected the FFAR2 protein with a molecular mass of about 50 kDa, which is comparable to the main band detected in pigs (Li et al., 2014) but different from the calculated size based on the sequence (Wang et al., 2009). The size difference can be explained, at least in part, by glycosylation sites, which are predicted by the NetNGlyc 1.0 server (http://www.cbs.dtu.dk, 2015/11/17), and the specificity was confirmed by the specific blocking peptide. The abundance of FFAR2 protein increased over time during the peripartal period. However, Friedrichs et al. (2014) did not find any changes in the expression of FFAR2 mRNA in the liver of dairy cows during the peripartal period. Bovine FFAR2 has a lower affinity for acetate and propionate than human FFAR2 (Hudson et al., 2012). Despite this, the concentration of propionate and butyrate in the portal vein (Lomax and Baird, 1983; Casse et al., 1994; Benson et al., 2002) reaches the half-maximal effective concentration (EC50) to activate the receptor, respectively. The portal vein concentrations indicate that the percentage of activation of this receptor in liver might be higher than its activation in other bovine tissues such as adipose tissue, because of lesser blood concentrations of butyrate and propionate in the periphery because of their percentage of hepatic extraction. However, the physiological significance of this receptor for liver metabolism has not been studied previously. Based on the number of animals in the current study, the correlation analysis may not be meaningful. However, it supports the decision to group by BHB level and reveals possibly contrasting associations of the two receptors with metabolic regulation during the peripartal period.