Characterization of the endocannabinoid system in subcutaneous adipose tissue in periparturient dairy cows and its association to metabolic profiles

Abstract

Adipose tissue (AT) plays a major role in metabolic adaptations in postpartum (PP) dairy cows. The endocannabinoid (eCB) system is a key regulator of metabolism and energy homeostasis; however, information about this system in ruminants is scarce. Therefore, this work aimed to assess the eCB system in subcutaneous AT, and to determine its relation to the metabolic profile in peripartum cows. Biopsies of AT were performed at 14 d prepartum, and 4 and 30 d PP from 18 multiparous peripartum cows. Cows were categorized retrospectively according to those with high body weight (BW) loss (HWL, 8.5 ± 1.7% BW loss) or low body weight loss (LWL, 2.9 ± 2.5% BW loss) during the first month PP. The HWL had higher plasma non-esterified fatty acids and a lower insulin/glucagon ratio PP than did LWL. Two-fold elevated AT levels of the main eCBs, N-arachidonoylethanolamine (AEA) and 2-arachidonoylglycerol (2-AG), were found 4 d PP compared with prepartum in HWL, but not in LWL cows. AT levels of the eCB-like molecules oleoylethanolamide, palmitoylethanolamide, and of arachidonic acid were elevated PP compared with prepartum in all cows. The abundance of monoglyceride lipase (MGLL), the 2-AG degrading enzyme, was lower in HWL vs. LWL AT PP. The relative gene expression of the cannabinoid receptors CNR1 and CNR2 in AT tended to be higher in HWL vs. LWL PP. Proteomic analysis of AT showed an enrichment of the inflammatory pathways' acute phase signaling and complement system in HWL vs. LWL cows PP. In summary, eCB levels in AT were elevated at the onset of lactation as part of the metabolic adaptations in PP dairy cows. Furthermore, activating the eCB system in AT is most likely associated with a metabolic response of greater BW loss, lipolysis, and AT inflammation in PP dairy cows.

Fig 1

Percentage of BW compared to week 1 postpartum (A) and the average BW (B) in HWL and LWL dairy cows. Cows were categorized either as high-weight loss (HWL, n = 9) or low-weight loss (LWL, n = 9), based on the percentage of BW loss between week 1 and 5 postpartum. Weekly averages of the percentage of BW compared to week 1 postpartum (A), and weekly averages of BW (B) are presented ± SEM. *P<0.05 in HWL compared to LWL; +P<0.1 in HWL compared to LWL cows.

Fig 2

Weekly average concentrations of plasma glucose (A), NEFA (B), insulin (C), and glucagon (D) in HWL and LWL cows. Cows were categorized either as high-weight loss (HWL, n = 9) or low-weight loss (LWL, n = 9), based on the percentage of BW loss between week 1 and 5 postpartum. Data represent the mean ± SEM. *P<0.05 in HWL compared to LWL; +P<0.1 in HWL compared to LWL cows.

Fig 3. Levels of AT eCBs that differed pre- and PP in the AT of dairy cows.

AT was sampled at 14 d prepartum (n = 10) and at 4 d PP (n = 10) for levels of AEA (A), 2-AG (B), OEA (C), PEA (D), and AA (E). Data represent the mean ± SEM.

Fig 4. Levels of eCBs in the AT of HWL and LWL dairy cows.

Cows were categorized as either high-weight loss (HWL, n = 9) or low-weight loss (LWL, n = 9), based on the percentage of BW loss between week 1 and 5 postpartum. Arachidonic acid (AA), 2-arachidonoylglycerol (2-AG), anandamide (AEA), oleoylethanolamide (OEA), and palmitoylethanolamide (PEA). Data represent the mean ± SEM. *P<0.05 vs. -14 days of the same group. #P<0.05 vs. +4 days of the LWL cows.

Fig 5. Longitudinal differential abundances of acute phase and oxidative stress-related proteins in AT of HWL and LWL cows according to proteomic analysis.

Cows were categorized either as high-weight loss (HWL, n = 9) or low-weight loss (LWL, n = 9) based on the percentage of BW loss between weeks 1 and 5 postpartum. The biopsies of subcutaneous AT from a subgroup of HWL and LWL AT (n = 5 per group) from 14 d prepartum, 4 d, and 30 d postpartum were analyzed by proteomic analysis. HP = haptoglobin; ORM1 = alpha-1-acid glycoprotein; VWF = von Willebrand factor; GSR = glutathione reductase.

Fig 6. Protein expression of PLIN1 and MGLL in ATs collected at 14 d prepartum and 4 d PP from HWL and LWL cows.

Cows were categorized as either high-weight loss (HWL, n = 9) or low-weight loss (LWL, n = 9) based on the percentage of BW loss between weeks 1 and 5 postpartum. The protein abundances of PLIN1 (A) and MGLL (B) were assessed by Western blotting analysis and corrected by β-actin as an internal standard. Samples for the western blot were prepared (×3.5), divided, and loaded to separate gels that ran simultaneously. The MGLL, PLIN1, and β-actin ran in parallel gels due to the proximity of the bands. Data represent the mean ± SEM. *P<0.05 in HWL vs. LWL AT prepartum. +P<0.1 in HWL vs. LWL AT postpartum.