The somatotropic axis during the physiological estrus cycle in dairy heifers—Effect on hepatic expression of GHR and SOCS2

Abstract

Pituitary growth hormone (GH) release and hepatic insulin-like growth factor-I (IGF-I) production increase after an injection of 17β-estradiol (E2) in ovariectomized dairy cattle. However, whether endogenous sexual steroid hormones also influence the hepatic GH receptor (GHR) signaling pathway during a physiological estrus cycle remains unclear. The aim of this study was to analyze the hepatic GHR signaling pathway during the luteal phase and after a period of increased E2 concentrations (after ovulation) as well as in 7 heifers before ovulation. Ovarian ultrasounds were performed daily during repeated physiological cycles (n = 56) of 30 Holstein Friesian heifers to determine ovulation [before ovulation (n = 7, bOv) and after ovulation 24-60 h after the appearance of estrus signs (n = 49, aOv)] and luteal phase (CLP; d 12 ± 1 after ovulation). Blood samples and liver biopsies were obtained, and blood concentrations of E2, P4, insulin-like growth factor (IGF)-I, IGF-II, and GH were measured. In the liver biopsies, we determined mRNA expression of the estrogen receptor α (ERα), GHR, Janus kinase 2 (JAK2), signal transducer and activator of transcription 5B (STAT5B), suppressor of cytokine signaling (SOCS)2 and 3, IGF-I, and IGF-II by quantitative reverse transcription-PCR. The concentration of E2 was higher bOv than aOv and CLP, as expected. The concentrations of IGF-I and GH were higher bOv and aOv compared with CLP. In contrast, concentrations of IGF-II were lower aOv compared with bOv and CLP. The mRNA expression of GHR was higher in liver biopsies obtained bOv compared with aOv and CLP. Notably, the expression of SOCS2 was higher bOv than aOv and in the CLP. Increased hepatic expression of SOCS2 during estrus was detectable when IGF-I concentrations were high; this result might indicate that SOCS2 expression attenuates the GHR signal transduction pathway during the phase of increased pituitary GH release. In conclusion, hepatic GHR and SOCS2 mRNA expression appeared to be promptly and sensitively regulated by increased E2 levels before ovulation of dairy heifers.

Serum IGFBP4 concentration decreased in dairy heifers towards day 18 of pregnancy

“…Before ET, the ovary of the recipient heifer was checked on day 6 by ultrasound for the development of a corpus luteum (CL), and the progesterone (P 4 ) blood concentration was quickly measured by use of the Immulite System (Siemens Healthcare Diagnostics, USA) (data not shown) as previously described [ 17 ]. If a CL with a diameter of > 1.5 cm was present and the P 4 serum concentration was > 1.0 ng/mL, the utilized male embryos (frozen in glycerol, stored at -196℃; produced and sexed by Masterrind) were thawed under sterile conditions using the Emcare Embryo Thawing System (ICPbio Reproduction, USA) according to the manufacturer's instructions.…”

“…Liver biopsies were obtained on day 18 as previously described [ 17 ]. Under sterile conditions, two or three liver biopsies, approximately 10 mm long and 3 mm in diameter, were collected into sterile Eppendorf cups, which were immediately frozen in liquid nitrogen and stored at -80℃ until mRNA extraction.…”

“…To determine the total serum IGF1 concentration, a commercial radioimmunoassay was used according to the manufacturer's instructions (IGF-1 IRMA A15729; Beckman Coulter, USA) and as previously reported [ 22 17 ]. The intra- and inter-assay CV results were 5.1 and 9.3%, respectively, while the lowest detection limit was 30 ng/mL.…”

Pregnancy‐Induced ISG‐15 and MX‐1 Gene Expression is Detected in the Liver of Holstein–Friesian Heifers During Late Peri‐Implantation Period

“…Non-pregnant heifers showed oestrous symptoms on day 18. Liver biopsies were taken on day 18 in pregnant or non-pregnant heifers as previously described (Mense et al 2015). Relative abundance of ISG-15 and MX-1 transcripts ( Table 1) was measured by real-time PCR as previously described (Mense et al 2015).…”

“…Liver biopsies were taken on day 18 in pregnant or non-pregnant heifers as previously described (Mense et al 2015). Relative abundance of ISG-15 and MX-1 transcripts ( Table 1) was measured by real-time PCR as previously described (Mense et al 2015). Statistical analyses were performed with open-source packages of the R project for Statistical Computing accessible throughout the Comprehensive R Archive Network (R Core Team 2013).…”