Characterization of ß-adrenergic receptors in bovine intramuscular and subcutaneous adipose tissue: comparison of lubabegron fumarate with ß-adrenergic receptor agonists and antagonists.

Chinese hamster ovary cell constructs expressing either the ß 1-, ß 2- or ß 3-adrenergic receptor (AR) were used to determine whether a novel ß-AR modulator, lubabegron fumarate (LUB; Experior, Elanco Animal Health) might exert greater potency for a specific ß-AR subtype. EC50 values calculated based on cAMP accumulation in dose response curves indicate that LUB is highly selective for the ß 3-AR subtype, with an EC50 of 6 × 10-9 M, with no detectible agonistic activity at the ß 2-AR. We hypothesized that the accumulation of lipolytic markers would reflect the agonist activity at each of the ß-receptor subtypes of the specific ligand; additionally, there would be differences in receptor subtype expression in subcutaneous (s.c.) and intrmuscular (i.m.) adipose tissues. Total RNA was extracted from adipose tissue samples and relative mRNA levels for ß 1-, ß2-, and ß 3-AR were measured using real-time quantitative polymerase chain reaction. Fresh s.c. and i.m. adipose tissue explants were incubated with isoproterenol hydrochloride (ISO; ß-AR pan-agonist), dobutamine hydrochloride (DOB; specific ß 1-AA), salbutamol sulfate (SAL; specific ß 2-AA), ractopamine hydrochloride (RAC), zilpaterol hydrochloride (ZIL), BRL-37344 (specific ß 3-agonist), or LUB for 30 min following preincubation with theophylline (inhibitor of phosphodiesterase). Relative mRNA amounts for ß 1-, ß 2-, and ß 3-AR were greater (P < 0.05) in s.c. than in i.m. adipose tissue. The most abundant ß-AR mRNA in both adipose tissues was the ß 2-AR (P < 0.05), with the ß 1- and ß 3-AR subtypes being minimally expressed in i.m. adipose tissue. ISO, RH, and ZH stimulated the release of glycerol and nonesterified fatty acid (NEFA) from s.c. adipose tissue, but these ß-AR ligands did not alter concentrations of these lipolytic markers in i.m. adipose tissue. LUB did not affect glycerol or NEFA concentrations in s.c. or i.m. adipose tissue, but attenuated (P < 0.05) the accumulation of cAMP mediated by the ß 1- and ß 2-AR ligands DOB and SAL in s.c. adipose tissue. Collectively, these data indicate that bovine i.m. adipose tissue is less responsive than s.c. adipose tissue to ß-adrenergic ligands, especially those that are agonists at the ß 1- and ß3-receptor subtypes. The minimal mRNA expression of the ß 1- and ß 3 subtypes in i.m. adipose tissue likely limits the response potential to agonists for these ß-AR subtypes.

Oleic acid in the absence of a PPARγ agonist increases adipogenic gene expression in bovine muscle satellite cells1.

We hypothesized that oleic acid (OA) in the absence of a thiazolidinedione (i.e., a synthetic peroxisome proliferator-activated receptorγ [PPARγ] agonist) would increase adipogenic gene expression in bovine muscle satellite cells (BSC). The BSC were cultured in differentiation medium containing 10 µM ciglitazone (CI), 100 µM OA, or 100 µM OA plus 10 µM CI (CI-OA). Control (CON) BSC were cultured only in differentiation media (containing 2% horse serum). The presence of myogenin, desmin, and paired box 7 proteins was confirmed in the BSC by immunofluorescence staining, demonstrating that we had isolated myogenic cells. The OA BSC had lesser paired box 3 (Pax3) and myogenic differentiation 1 expression but greater Pax7 and mygogenin (MYOG) expression (P < 0.05), than the CON BSC. The CI BSC had greater Pax3, Pax7, and MYOG expression than CON BSC (P < 0.05), suggesting that CI would promote BSC myogenesis under pro-myogenic conditions (i.e., when cultured with horse serum). However, both the OA and CI treatments upregulated the expression of PPARγ, CCAAT/enhancer-binding protein alpha (C/EBPα) and C/EBPß, sterol regulatory element-binding protein 1, lipoprotein lipase, and glycerol-3-phosphate acyltransferase 3 gene expression, as well as media adiponectin concentration (P < 0.05). The CI, OA, and CI-OA treatments also increased triacylglycerol and lipid droplet accumulation, in spite of upregulation (relative to CON BSC) of adenosine monophosphate-activated protein kinase alpha-1, perilipin 2 (PLIN2), and PLIN3 in BSC and downregulation of G protein-coupled protein receptor 43, acyl-CoA synthetase long chain family member 3, and stearoyl-CoA desaturase (P < 0.05). These results indicate that OA in the absence of a synthetic PPARγ agonist can effectively increase adipogenic gene expression in BSC.

Adipogenic/lipogenic gene expression and fatty acid composition in chuck, loin, and round muscles in response to grain feeding of Yanbian Yellow cattle.

We hypothesized that gene expression and fatty acid composition would differ among different muscle depots and over time on a grain-based finishing diet. Additionally, we hypothesized that the concentration of SFA would decrease and the concentration of MUFA would increase proportionately with increases in percent intramuscular lipid (%IML). Ten Yanbian Yellow cattle steers (approximately 8 mo of age) were fed a corn-based diet in which the proportion of corn in the ration was increased at 4-mo intervals. Muscle samples were taken by biopsy from the chuck (trapezius), loin (longissimus dorsi), and round (biceps femoris) muscles at 12, 16, 20, 24, and 28 mo of age. The %IML increased from 12 to 28 mo of age, especially between 24 and 28 mo of age, with loin > round > chuck (age × muscle interaction P < 0.001). The percentage (g/100 g total fatty acids) of oleic acid (18:1n-9), linoleic acid (18:2n-6), and α-linolenic acid (18:3n-3), and the MUFA:SFA ratio increased with age, whereas palmitic (16:0) and stearic acid (18:0) decreased with age in all muscles (age effect P < 0.001). The expression of sterol regulatory element binding protein (SREBP1), adipose tissue fatty acid- binding protein (FABP4), stearoyl-CoA desaturase (SCD), acetyl-CoA carboxylase (ACC1), and lipoprotein lipase (LPL) increased, whereas the expression of peroxisome proliferator-activated receptor gamma (PPARγ) and fatty acid synthase (FASN) decreased with age. Expression of PPARγ, FABP4, SREBP1, SCD, FASN, ACC1, and LPL was greater in the loin than in the chuck or round (age × muscle interaction P < 0.001), although the MUFA:SFA ratio was greater in the chuck than in the loin or round (muscle effect P < 0.001). In conclusion, adipogenic gene expression was greater in the loin than in the chuck or round muscles, consistent with the greater %IML of the loin. However, the greater SCD gene expression in the loin did not result in a greater amount of MUFA in the loin, relative to the chuck and round.

18F-FDG uptake for prediction EGFR mutation status in non-small cell lung cancer.

Epidermal growth factor receptor (EGFR) mutations in non-small cell lung cancer (NSCLC) are a response to EGFR-tyrosine kinase inhibitor. However, a lack of sufficient tumor tissue has been a limitation for determining EGFR mutation status in clinical practice. The objective of this study was to predict EGFR mutation status in NSCLC patients based on a model including maximum standardized uptake value (SUVmax) and clinical features.We retrospectively reviewed NSCLC patients undergoing EGFR mutation testing and pretreatment positron emission tomography/computed tomography between March 2009 and December 2013. The relationships of EGFR mutations with both SUVmax and patient characteristics were evaluated, and a multivariate logistic regression analysis was performed. The model was assessed by area under the receiver-operating characteristic curve (AUC) and was prospectively validated during January to June 2014.Three hundred and sixteen patients meeting the criteria were enrolled for model construction. The SUVmax values were significantly lower for EGFR mutations (mean, 9.5 ±â€Š5.74) than for EGFR wild-type (mean, 12.7 ±â€Š6.43; P < 0.001). ROC curve analysis showed that the SUVmax cutoff point was 8.1, for which the AUC was 0.65 (95% confidence interval [CI], 0.60-0.72). In addition, multivariate analysis also showed that low SUVmax (≤8.1) was a predictor of EGFR mutations, for which the AUC was 0.77, combining nonsmoking history and primary tumor size (≤5 cm). Eighty-five patients were enrolled to validate the predictive model, and the overall accuracy, sensitivity, and specificity were 77.6%, 64.6% (95% CI 40.7-82.8), and 82.5% (95% CI 70.9-91.0), respectively.The specific FDG uptake value could be considered to effectively predict EGFR mutation status of NSCLC patients by considering smoking history and primary tumor size when genetic tests are not available.

Effects of dietary linseed oil and propionate precursors on ruminal microbial community, composition, and diversity in Yanbian yellow cattle.

The rumen microbial ecosystem is a complex system where rumen fermentation processes involve interactions among microorganisms. There are important relationships between diet and the ruminal bacterial composition. Thus, we investigated the ruminal fermentation characteristics and compared ruminal bacterial communities using tag amplicon pyrosequencing analysis in Yanbian yellow steers, which were fed linseed oil (LO) and propionate precursors. We used eight ruminally cannulated Yanbian yellow steers (510 ± 5.8 kg) in a replicated 4 × 4 Latin square design with four dietary treatments. Steers were fed a basal diet that comprised 80% concentrate and 20% rice straw (DM basis, CON). The CON diet was supplemented with LO at 4%. The LO diet was also supplemented with 2% dl-malate or 2% fumarate as ruminal precursors of propionate. Dietary supplementation with LO and propionate precursors increased ruminal pH, total volatile fatty acid concentrations, and the molar proportion of propionate. The most abundant bacterial operational taxonomic units in the rumen were related to dietary treatments. Bacteroidetes dominated the ruminal bacterial community and the genus Prevotella was highly represented when steers were fed LO plus propionate precursors. However, with the CON and LO diet plus malate or fumarate, Firmicutes was the most abundant phylum and the genus Ruminococcus was predominant. In summary, supplementing the diets of ruminants with a moderate level of LO plus propionate precursors modified the ruminal fermentation pattern. The most positive responses to LO and propionate precursors supplementation were in the phyla Bacteriodetes and Firmicutes, and in the genus Ruminococcus and Prevotella. Thus, diets containing LO plus malate or fumarate have significant effects on the composition of the rumen microbial community.

Conjugated fatty acids and methane production by rumen microbes when incubated with linseed oil alone or mixed with fish oil and/or malate.

We hypothesized that manipulating metabolism with fish oil and malate as a hydrogen acceptor would affect the biohydrogenation process of α-linolenic acid by rumen microbes. This study was to examine the effect of fish oil and/or malate on the production of conjugated fatty acids and methane (CH4 ) by rumen microbes when incubated with linseed oil. Linseed oil (LO), LO with fish oil (LO-FO), LO with malate (LO-MA), or LO with fish oil and malate (LO-FO-MA) was added to diluted rumen fluid, respectively. The LO-MA and LO-FO-MA increased pH and propionate concentration compared to the other treatments. LO-MA and LO-FO-MA reduced CH4 production compared to LO. LO-MA and LO-FO-MA increased the contents of c9,t11-conjugated linoleic acid (CLA) and c9,t11,c15-conjugated linolenic acid (CLnA) compared to LO. The content of malate was rapidly reduced while that of lactate was reduced in LO-MA and LO-FO-MA from 3 h incubation time. The fold change of the quantity of methanogen related to total bacteria was decreased at both 3 h and 6 h incubation times in all treatments compared to the control. Overall data indicate that supplementation of combined malate and/or fish oil when incubated with linseed oil, could depress methane generation and increase production of propionate, CLA and CLnA under the conditions of the current in vitro study.

Rumen microbial response in production of CLA and methane to safflower oil in association with fish oil or/and fumarate.

Supplementation effect of fish oil and/or fumarate on production of conjugated linoleic acid (CLA) and methane by rumen microbes was examined when incubated with safflower oil. One hundred and twenty milligrams of safflower oil (SO), safflower oil with 24 mg fish oil (SOFO), safflower oil with 24 mmol/L fumarate (SOFA), or safflower oil with 24 mg fish oil and 24 mmol/L fumarate (SOFOFA) were added to the 90 mL culture solution. The culture solution was also made without any supplements (control). The SOFA and SOFOFA increased pH and propionate (C3) compared to other treatments from 3 h incubation time. An accumulated amount of total methane (CH(4) ) for 12 h incubation was decreased by all the supplements compared to control. The concentrations of c9,t11CLA for all the incubation times were increased in the treatments of SOFO, SOFA and SOFOFA compared to SO. The highest concentration of c9,t11CLA was observed from SOFOFA among all the treatments at all incubation times. Overall data indicate that supplementation of combined fumarate and/or fish oil when incubated with safflower oil could depress CH(4) generation and increase production of C(3) and CLA under the condition of current in vitro study.