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* Division of Animal Sciences, University of Missouri, Columbia 65211
Department of Animal Science, University of Minnesota, St. Paul 55108
2 Corresponding author: lucym{at}missouri.edu
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ABSTRACT |
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 TOP ABSTRACT INTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONCONCLUSIONSACKNOWLEDGEMENTSREFERENCES |
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Key Words: growth hormone receptor • liver • postpartum • nutrition
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INTRODUCTION |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONCONCLUSIONSACKNOWLEDGEMENTSREFERENCES |
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The liver-specific GHR 1A mRNA is believed to give rise to the bulk of the functionally active GHR in liver (Radcliff et al., 2003a). Binding of GH to the GHR causes a variety of physiological responses, one of which is the induction of IGF-I gene expression via a distal Stat5b binding site (Wang and Jiang, 2005). The GH-induced synthesis and secretion of IGF-I by liver creates an endocrine feedback loop that involves IGF-I feeding back negatively on the hypothalamus and pituitary to control GH secretion (LeRoith et al., 2001). In dairy cows, the GHR 1A mRNA decreases shortly before calving and remains low for about 1 wk (Kobayashi et al., 1999; Wook Kim et al., 2004). The mechanisms that cause the precipitous decline in GHR 1A before calving are unknown, but seem to be specific to dairy cows because the change in GHR 1A does not occur in beef cows (Jiang et al., 2005). The decrease in GHR 1A is believed to mediate the uncoupling of the GH axis that typically occurs in postpartum dairy cows (Lucy et al., 2001). One hypothesis is that the uncoupling of the axis (loss of GHR expression) decreases blood IGF-I concentrations and relieves negative feedback. Loss of negative feedback increases postpartum GH concentrations.
Blood GH concentrations remain elevated for several weeks after calving (Radcliff et al., 2003b). Increase in blood GH coordinates nutrient partitioning by increasing adipose tissue mobilization (NEFA release) and by increasing the gluconeogenic rate in liver (Etherton and Bauman, 1998). Expression of GHR 1A increases postpartum and the GH axis is recoupled. Mechanisms that underlie the increase in GHR 1A after calving are unknown. Infusion of insulin into postpartum dairy cows increased liver GHR 1A and IGF-I (Butler et al., 2003; Rhoads et al., 2004). The possibility that insulin controls recrudescence of GHR 1A (and hence the re-coupling of the GH axis) is consistent with the stimulatory effects of insulin on the liver GHR (Bereket et al., 1999; Lucy, 2004).
Postpartum feeding affects GH axis recoupling (McGuire et al., 1992), perhaps through its effects on insulin and the GHR. We hypothesized that postpartum DMI would control the GHR 1A mRNA in a manner that was consistent with an intake-dependent recoupling process. In this study, DMI was limited (restricted) in a group of postpartum dairy cows. The GHR 1A, total GHR (GHRtot; all GHR mRNA variants), IGF-I, and cyclophilin mRNA expression within restricted cows was compared with that of cows fed ad libitum (control). Production characteristics and plasma hormones and metabolites were also compared to evaluate the capacity for GH axis recoupling within each group of cows.
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MATERIALS AND METHODS |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONCONCLUSIONSACKNOWLEDGEMENTSREFERENCES |
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Liver Biopsies
Liver biopsies (approximately 100 mg) were collected from each cow 14 d before expected calving (actual day relative to calving was –10.0 ± 1.4 d), and then on d 1, 7, 14, and 21 after calving. Biopsies were done in the afternoon before afternoon feeding. Biopsies were collected with a 14-gauge needle (Radcliff et al., 2003b). Samples were immediately frozen in liquid nitrogen, brought to the laboratory, and stored at –80°C until analysis.
Blood Samples
A single blood sample was collected immediately before each biopsy. Additional blood samples were collected daily from calving (d 0) through d 14. Additional samples were collected at about the same time as the biopsy samples (in the afternoon before the afternoon feeding). Evacuated glass tubes containing EDTA (Vacutainer, Becton Dickinson and Co., Franklin Lakes, NJ) were used and blood samples were collected from the coccygeal vein. Blood samples were stored on ice for transport to the laboratory where they were centrifuged at 1,200 x g for 20 min to harvest plasma. Plasma was transferred to a polypropylene tube and frozen at – 20°C until assayed.
Hormone and Metabolite Analyses
Plasma glucose, NEFA, and BHBA were quantified in samples collected prepartum and in samples collected on d 1, 2, 4, 6, 8, 10, 12, 14, and 21 postpartum. Commercially available colorimetric assay kits were used [glucose (ThermoDMA, Louisville, CO); NEFA (Wako Pure Chemical Industries Ltd., Osaka, Japan); and BHBA (Randox Laboratories, San Francisco, CA)]. Intraassay CV were 4.7, 2.6, and 4.8%, respectively.
Plasma GH and IGF-I concentrations were quantified using radioimmunoassays previously described (Radcliff et al., 2003b). The intra- and interassay CV were 5.0 and 2.0% for GH, and 5.3 and 8.7% for IGF-I, respectively. Plasma insulin concentrations were quantified using a specific, double-antibody, equilibrium radioimmunoassay (Elsasser et al., 1986) with some modifications described as follows. Preparation of bovine insulin (Sigma Chemical Co., St. Louis, MO) for iodination and for standard curve material followed the method of Sodoyez et al. (1975) for preparation of zinc-free insulin. Briefly, 100 mg of bovine insulin was dissolved in 25 mL of 0.01 N HCl and zinc ions were chelated by the addition of 1 mL of 50 mM EDTA. Subsequently, zinc-free insulin was then precipitated from the solution by the addition of 13 mL of 0.2 M sodium citrate (pH 5.6) at room temperature and the precipitate was recovered by centrifugation at 4,000 x g for 10 min and then lyophilized. Ten micrograms of zinc-free bovine insulin was then solubilized in 50 µL of water, combined with 500 µCi 125I-Na, and incubated in the presence of 100 µg of Iodogen (Pierce Chemical Co., Rockford, IL) for 6 min with gentle mixing. Recovery of the monoiodinated form of 125I-bovine insulin was achieved by differential elution from a 10-mL Sep-Pak C18 Cartridge (Waters Corp., Milford, MA) as previously described by Deleo (1994). Briefly, the Sep-Pak C18 cartridge was initially washed with 10 mL of 50% (vol/vol) acetonitrile containing 50 mM triethylamine solution (adjusted to pH = 3 with phosphoric acid), followed by 10 mL of deionized water before addition of the iodination mixture. The cartridge was then washed sequentially with 5 mL of 0.4 M phosphate buffer (pH = 7.4), 10 mL of 29% (vol/vol) acetonitrile containing 50 mM triethylamine, 5 mL of 10% (vol/vol) acetonitrile containing 0.2 M ammonium acetate (pH = 5.5), and finally 5 mL of 50% (vol/vol) acetonitrile containing 0.2 M ammonium acetate (pH = 5.5). This final fraction was collected and diluted to 25,000 cpm per 100 µL of assay buffer (PABET; consisting of 0.1% gelatin, 0.01 M EDTA, 0.9% NaCl, 0.01 M PO4, 0.01% sodium azide, 0.1% Tween-20, pH = 7.1). Guinea pig antibovine insulin antiserum (generously provided by Ted Elsasser, USDA ARS Growth Biology Lab, Beltsville, MD; Elsasser et al., 1986) was diluted to a final tube dilution of 1:167,000 in PABET. Standard concentrations of zinc-free bovine insulin (0.064 to 40 ng/tube) and increasing volumes of a bovine plasma pool (25 to 300 µL) were added to assay tubes in quadruplicate and the total volume balanced to 300 µL/tube with PABET. All plasma samples (100-µL aliquots) to be analyzed were assayed in triplicate. All components were then incubated at 4°C for 24 h. The antigen-antibody complex was precipitated following a 15-min incubation at 22°C with 100 µL of a pre-precipitated sheep-anti-guinea pig second antibody. The second antibody complex was then precipitated by centrifugation at 3,000 x g for 30 min, and the supernatant removed by aspiration. Assay tubes containing the precipitate were counted for 1 min in a gamma counter. Standards and plasma aliquots of the bovine plasma pool were linear (log/logit transformation; R2 > 0.98) and parallel over a mass of 0.064 to 40 ng/tube and a plasma volume of 25 to 300 µL. Total specific binding was 38%, the minimum detectable concentration was 0.064 ng/tube, percentage recovery of mass was > 98 %, and the inter- and intraassay coefficients of variations were less than 10%.
BW and Milk Production
Body weight was measured immediately before each biopsy. Cows were milked twice daily and milk weight was measured electronically. Daily milk yield (sum of a.m. and p.m. milk weights) was calculated for the first 21 d of lactation.
RNA Isolation and Quantitative Real-Time PCR
Total cellular RNA was isolated from each liver sample using the TRIzol procedure (Life Technologies Inc., Gaithersburg, MD) and dissolved in 50 µL of diethyl pyrocarbonate-treated water. Concentrations of RNA were determined by measuring absorbance at 260 nm. The purity of the RNA was determined by calculating the ratio of absorbances at 260 and 280 nm and by electrophoresis of an RNA aliquot (2.5 µg) from each sample through a 1% agarose gel in Tris-borate/EDTA buffer (0.09 M Tris-borate and 0.002 M EDTA) with ethidium bromide (0.5 µg/mL). All samples had intact 28S and 18S ribosomal RNA and were used for subsequent analyses. Isolated RNA was stored at –80°C until analysis.
Quantitative real-time reverse transcription PCR was used to determine the amounts of GHR 1A, GHRtot, IGF-I, and cyclophilin mRNA as previously described (Radcliff et al., 2003b). Total cellular RNA (2.1 µg) was used for cDNA synthesis using the Gib-coBRL Superscript First-Strand RT-PCR kit (Life Technologies Inc.). Amplification mixes (25 µL) were prepared according to the manufacturer’s instructions and contained 5 µL of cDNA (25 ng), 500 nM of forward primer (2.5 µL), 500 nM of reverse primer (2.5 µL), 100 nM of probe (0.025 µL), 12.5 µL of Brilliant Plus Quantitative PCR reagent mixture (Stratagene; La Jolla, CA), and 2.5 µL of RNase-free distilled water. Each PCR plate contained a no-template control, internal controls (low, medium, and high), and RNA standards. Reactions were run in triplicate and fluorescence quantified with the ABI PRISM 7700 Sequence Detector (Applied Biosystems, Foster City, CA). Analyses of amplification plots were done using the Sequence Detection Software of Applied Biosystems. Standard curves were subjected to linear regression and used to estimate femtograms of target RNA in the sample. The final values are reported as femtograms of target RNA per 25 ng of total cellular RNA in the reverse transcription (RT) reaction.
Statistical Analyses
Data collected during the treatment period (d 1 to 14 after calving) were analyzed as repeated measures using the Proc Mixed procedures of SAS (SAS Inst., Inc., Cary, NC). The model for all analyses included the main effects of treatment (feed-restricted or control), day after calving, and treatment x day interaction. The appropriate covariance structure for each variable was determined by testing several different covariance structures and then choosing the covariance structure with the lowest fit statistics. The covariance structure used for BW analyses was compound symmetry. The covariance structure used for the GHR 1A mRNA, GHRtot mRNA, IGF-I mRNA, and hormones and metabolites was heterogeneous compound symmetry. The covariance structure used for cyclophilin mRNA was unstructured. Milk yield was analyzed using autoregressive and DMI was analyzed using heterogeneous autoregressive covariance structures. A test for heterogeneity of regression was used to estimate the effect of treatment on the increase in milk production and GHR 1A after calving. Data collected before calving (prepartum sample) and after the feed restriction (d 21 postpartum) were analyzed for the effect of treatment on the individual day (prepartum sample or d 21 postpartum). Plasma hormone and metabolite concentrations and the amount of mRNA were compared for the prepartum sample and the sample collected on d 1 postpartum to determine if the concentration or amount changed after calving. A Type I error rate of P > 0.10 was considered nonsignificant unless stated otherwise.
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RESULTS |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONCONCLUSIONSACKNOWLEDGEMENTSREFERENCES |
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). Feed-restricted cows consumed 65% of the DM relative to controls. Body weight was similar before calving for feed-restricted and control cows (mean = 746 ± 19 kg) and decreased after calving (day effect; P < 0.01). Decrease in BW during the treatment period was greater in feed-restricted than in control cows; BW decreased from 653 ± 27 kg (d 1) to 590 ± 27 kg (d 14) for feed-restricted and from 683 ± 29 (d 1) to 661 ± 29 (d 14) for control cows (treatment x day; P = 0.02). Feed-restricted and control cows had similar BW on d 21 (636 ± 19 kg). An effect of day (P < 0.01) was detected on milk production because daily milk production increased from d 1 to 14 (Figure 1B). When analyzed across d 1 to 14 with day as a discrete variable, feed-restricted and control cows were similar for milk production (effect of treatment; P = 0.12). Analyses of milk production with day as a continuous variable by heterogeneity of regression demonstrated that rate of increase in postpartum milk production was less for feed-restricted than control cows (treatment x day; P < 0.001). Milk production was similar for feed-restricted and control cows on d 21 postpartum (40 ± 3 kg/d).
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). Postpartum plasma NEFA concentrations were greater (P < 0.05) in feed-restricted than control cows during the feed-restriction period. A modest decline in plasma NEFA occurred over time in both groups (day effect; P < 0.05). Plasma NEFA concentrations were similar for feed-restricted and control cows on d 21 postpartum (7 d after the end of feed restriction; 392 ± 59 µEq/L).
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). Effects of day (P < 0.001) and a treatment x day interaction (P < 0.007) were detected on plasma BHBA concentrations during the feed restriction. Plasma BHBA concentrations increased in feed-restricted cows and were generally greater in feed-restricted cows after d 2 postpartum. Plasma BHBA concentrations were similar for feed-restricted and control cows on d 21 postpartum (1.19 ± 0.06 mmol/L).
Plasma glucose concentrations were similar for feed-restricted and control cows before calving (73 ± 16 mg/dL). Relative to prepartum values, plasma glucose concentrations were decreased (P < 0.05) by d 1 after calving (Figure 2C). A tendency occurred for feed-restricted cows to have reduced (P = 0.11) plasma glucose during feed restriction. An effect of day (P < 0.01) was detected for postpartum plasma glucose because plasma glucose concentrations decreased for approximately 1 wk postpartum and then subsequently increased. Plasma glucose concentrations were similar for feed-restricted and control cows on d 21 postpartum (66 ± 3 mg/dL).
Plasma GH, IGF-I, and Insulin
Plasma GH concentrations were similar for feed-restricted and control cows before calving (4.2 ± 0.4 ng/mL) and did not increase by d 1 after calving. After calving, plasma GH concentrations tended (P = 0.11) to be greater for feed-restricted compared with control cows during feed restriction (Figure 3A). No effect of day or treatment x day was detected for plasma GH. Plasma GH concentrations did not differ between feed-restricted and control cows on d 21 postpartum (5.4 ± 0.7 ng/mL).
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). Plasma IGF-I concentrations were generally greater in control cows after d 2 postpartum. Plasma IGF-I concentrations increased from d 4 to 14 in feed-restricted and control cows. Greater IGF-I in control cows persisted for at least 7 d after the end of feed restriction because plasma IGF-I was greater (P < 0.07) in control than feed-restricted cows on d 21 postpartum (94.0 ± 5.3 and 79.2 ± 4.8 ng/mL, respectively).
Plasma insulin concentrations were similar for feed-restricted and control cows before calving (5.9 ± 0.8 ng/mL). Relative to prepartum concentrations, plasma insulin decreased (P < 0.001) by d 1 after calving (Figure 3C). Plasma insulin concentrations were similar for feed-restricted and control cows during feed restriction. An effect of day (P < 0.002) was detected on postpartum plasma insulin concentrations because plasma insulin concentrations increased postpartum. Plasma insulin concentrations were similar for feed-restricted and control cows on d 21 postpartum (4.3 ± 0.3 ng/mL).
GHR 1A, IGF-I, GHRtot, and Cyclophilin mRNA in Liver
Amount of cyclophilin mRNA did not differ between feed-restricted and control cows before calving (434 ± 83 fg/25 ng RT, where 25 ng RT is the amount of cellular RNA in the RT reaction) but subsequently increased (P < 0.01) after calving (d 1; 841 ± 99 fg/25 ng RT). No effects of treatment, day, or treatment by day interaction were detected for cyclophilin mRNA during feed restriction. Cyclophilin mRNA was similar for feed-restricted and control cows on d 21 (1035 ± 214 fg/25 ng RT).
Amount of GHR 1A mRNA was similar for feed-restricted and control cows before calving (1,047 ± 223 fg/25 ng RT; Figure 4A). After calving (d 1), amount of GHR 1A mRNA decreased (123 ± 28 fg/25 ng RT; P < 0.01). There was an effect of day (P < 0.01) on GHR 1A mRNA during feed restriction because the amount of GHR 1A mRNA increased in both feed-restricted and control cows from d 1 to 14. The treatment x day interaction approached significance (P = 0.11). Analyses of GHR 1A mRNA with day as a continuous variable by heterogeneity of regression demonstrated that rate of increase in postpartum GHR 1A was less (P < 0.01) for feed-restricted than for control cows (treatment x day;). On d 21 (7 d after the end of feed restriction), the amount of GHR 1A mRNA was greater (P < 0.01) in control than in feed-restricted cows. Changes in GHR 1A mRNA were not mirrored by GHRtot because the amount of GHRtot increased from prepartum (506 ± 96 fg/25 ng RT) to d 1 (874 ± 63 fg/25 ng RT). No effects of treatment, day, or treatment by day were detected for GHRtot during feed restriction. Amount of GHRtot was similar for feed-restricted and control cows on d 21 (834 ± 178 fg/25 ng RT).
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). After calving (d 1), amount of IGF-I mRNA (39 ± 8 fg/25 ng RT) decreased (P < 0.05). There was an effect of day (P < 0.05) on for IGF-I mRNA during the feed restriction period because the amount of IGF-I mRNA increased in both feed-restricted and control cows from d 1 to 14. No effects of treatment or treatment x day interaction were detected for amounts of IGF-I mRNA. Amount of IGF-I mRNA was similar for feed-restricted and control cows on d 21 (258 ± 94 fg/25 ng RT). |
DISCUSSION |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONCONCLUSIONSACKNOWLEDGEMENTSREFERENCES |
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) and increased plasma NEFA and BHBA (Figure 2). Increase in plasma NEFA and BHBA indicated that lipid mobilization was increased in response to feed restriction. As expected, the GHR 1A mRNA was elevated in prepartum samples and decreased by d 1 postpartum (Figure 4A). This distinctive decrease in GHR 1A mRNA was observed in our previous studies (Kobayashi et al., 1999; Radcliff et al., 2003b). Amount of GHR 1A increased postpartum, but was less for feed-restricted cows than for control cows. A distinct difference in GHR 1A mRNA was present 7 d after the end of feed restriction (d 21). Thus, postpartum DMI can affect GHR 1A mRNA expression and a carryover effect seems to persist for at least 1 wk. This study was the latest in a series of experiments in which the relationship between nutrient intake and GHR expression was examined. We subjected later-lactation dairy cattle (153 to 265 DIM) to feed restriction in a manner consistent with periparturient intake (gradual decrease to 25% intake) and found that IGF-I mRNA decreased, but GHR 1A remained relatively stable (Kobayashi et al., 2002). A 72-h fast decreased both GHR 1A and IGF-I mRNA, but had little effect on GHR 1B (alternative GHR variant; Wang et al., 2003). The feed restriction we imposed was relatively modest (approximately 70%). This reduction in intake reduced the postpartum increase in GHR 1A through at least 21 DIM.
Liver GHR concentration and activity depend on nutritional status with improved nutritional status associated with greater GHR expression and activity (McGuire et al., 1992; Beauloye et al., 2002). The link between nutrition and liver GHR expression may depend on blood insulin concentration. Indeed, insulin increased GHR expression in a variety of physiological settings (Lucy, 2004). Insulin infusion into postpartum dairy cows increased liver GHR 1A and IGF-I mRNA (Butler et al., 2003; Rhoads et al., 2004). We measured plasma insulin in the present study because we hypothesized that feed restriction would decrease blood insulin concentration and correlate with the observed decrease in GHR 1A. Feed-restricted cows did not have reduced blood insulin concentration (Figure 3C) despite causing reduced blood glucose (Figure 2C). Our single daily blood sample taken from a peripheral site may have been inadequate to capture the true daily insulin status as manifested in the portal vein. It is also possible that insulin may not drive the observed increase in GHR 1A during the early postpartum period; other metabolic signals may be involved (Brameld et al., 1999).
The GHR 1A mRNA is a liver-specific GHR mRNA variant (Jiang and Lucy, 2001). The amount of GHR 1A mRNA is correlated with GH binding in liver (Radcliff et al., 2003a). Binding of GH to the liver GHR causes activation of the Jak/stat5b pathway (Herrington and Carter-Su, 2001) that impinges upon a distal stat5b binding site within the IGF-I gene (Wang and Jiang, 2005). Activation of IGF-I gene expression leads to liver IGF-I synthesis and secretion. Changes in GHR 1A mRNA, therefore, are generally associated with changes in liver IGF-I mRNA and blood IGF-I concentrations (Radcliff et al., 2003b). The amount of GHR 1A increased postpartum and feed restriction slowed the increase in GHR 1A. The amount of IGF-I mRNA increased postpartum as well, but no effect of feed restriction was detected for IGF-I mRNA and the effects of feed restriction on plasma IGF-I were modest. The postpartum increase in liver IGF-I mRNA and in blood IGF-I concentrations may be controlled partially by mechanisms that are independent of postpartum nutrition. These nutritionally independent mechanisms may not depend on GHR 1A expression.
Our model for postpartum GH incorporates a negative feedback effect for IGF-I on GH. Presence of this specific feedback loop is well supported by lines of evidence arising from diverse species (LeRoith et al., 2001). We think that the decrease in GHR 1A caused a decrease in IGF-I mRNA and in blood IGF-I. The decrease in blood IGF-I leads to a loss of negative feedback and greater blood GH. In this study, the GHR 1A and IGF-I mRNA decreased after calving (Figure 4). Plasma IGF-I also decreased after calving and tended to remain lower in feed-restricted cows (Figure 3B). Growth hormone concentrations tended to increase in feed-restricted cows relative to control, but the response was transitory (primarily occurring during the first week). Blood GH concentrations were similar for feed restricted and control on d 21 when GHR 1A mRNA and blood IGF-I were reduced for feed-restricted cows.
Growth hormone is released episodically (Breier et al., 1986). A single sample per day and the limited number of experimental cows may not have provided a true reflection of the effects of feed restriction on circulating GH concentrations. It is also possible that the control of GH secretion in early postpartum cows is more complex and involves a variety of factors, one of which is blood IGF-I concentration that is controlled, in part, by GHR 1A. Other components of the axis act to control blood GH concentrations as well. This could include specific nutritional cues that act directly on the hypothalamus and pituitary (Scacchi et al., 2003) and on GH and IGF-binding proteins whose concentrations change dynamically during the postpartum period (Vicini et al., 1991). Sequestration of GH or IGF-I by binding proteins could change endocrine feedback signals and modify the systemic control of GH.
We measured cyclophilin because we originally believed that it would be a suitable housekeeping gene for the mRNA that we studied. We found that the amount of cyclophilin mRNA increased postpartum. Similar results were reported by other laboratories (Rhoads et al., 2003) and from work done within our laboratory (Radcliff et al., 2003b). Thus, the present study represents the third study to show that cyclophilin mRNA is a poor candidate for a housekeeping gene for periparturient liver because the amount of cyclophilin mRNA increases postpartum. Apparently, cyclophilin mRNA responds to a variety of metabolic signals (Rhoads et al., 2003). Choosing an appropriate housekeeping gene is difficult (Bustin, 2002), particularly when testing samples of pre- and postpartum liver. We chose not to pursue additional housekeeping genes and instead presented the results as a function of the starting RNA amount in the reverse transcription. Cyclophilin increased postpartum, but treatment did not affect amount of cyclophilin mRNA. We can infer from these results, therefore, that a treatment bias did not exist for the total mRNA or total cDNA content within the samples. The latter result was expected because an equivalent amount of RNA was included in each reverse transcription reaction.
The GHRtot mRNA (a measure of all GHR mRNA variants) actually increased in the present study when GHR 1A mRNA decreased. We typically observe a non-significant decrease in GHRtot at calving and we were surprised to find the increase in GHRtot mRNA. Composition of liver GHR mRNA is diverse and some GHR mRNA transcripts may increase when GHR 1A mRNA decreases. We do not fully understand these global mRNA changes that potentially influence GHRtot. Our previous work showed that liver GHR protein was highly correlated with GHR 1A mRNA (Radcliff et al., 2003a). This high correlation may reflect the highly efficient translation of GHR 1A mRNA relative to other GHR mRNA variants (Jiang and Lucy, 2001). We predict, therefore, that although GHRtot mRNA increased, the decrease in GHR 1A mRNA led to a decrease in GHR protein in feed-restricted cows.
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CONCLUSIONS |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONCONCLUSIONSACKNOWLEDGEMENTSREFERENCES |
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ACKNOWLEDGEMENTS |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONCONCLUSIONSACKNOWLEDGEMENTSREFERENCES |
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FOOTNOTES |
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Received for publication June 3, 2005. Accepted for publication October 3, 2005.
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONCONCLUSIONSACKNOWLEDGEMENTSREFERENCES |
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; n = 6) or fed for ad libitum consumption (•; n = 5) from d 0 to 14 postpartum. Data are least squares means ± SEM. The DMI could not be collected for d 14 postpartum because cows returned to the milking herd (group feeding) on the morning of d 14.
