Association of leptin genotype with growth performance, adipocyte cellularity, meat quality, and fatty acid profile in beef steers fed flaxseed or high-oleate sunflower seed diets with or without triticale dried distiller’s grains

Abstract

Leptin genotypes can be identified as homozygous normal (CC), homozygous mutant (TT), and heterozygous (CT) based on a single-nucleotide polymorphism in exon 2 of the leptin gene, which has been associated with feed intake and fat deposition in cattle. The experiment was designed as 2 × 2 × 2 factorial with three main factors: (1) genotype (CT or TT) and diets fed 2) with or without triticale dried distiller’s grains with solubles (DDG), and 3) with either flaxseed (FS) or high-oleate sunflower seed (SS). Evaluations included growth performance, subcutaneous fat deposition, adipocyte cellularity, meat quality, and fatty acid (FA) profile of various depots. Beef steers (n = 40, 459 ± 31 kg) of either CT or TT genotypes were housed in individual pens with ad libitum access to one of the four diets: 75% steam-rolled barley + 10% barley silage with 10% FS or SS (non-DDG diets, NDG) and 46.5% barley + 10% barley silage + 30% DDG, with 8.5% FS or SS, all on a dry matter basis. Growth performance, ultrasound subcutaneous fat thickness, rib eye area (REA), and plasma FA were measured prior to and during the finishing period. At slaughter, samples of subcutaneous fat, perirenal fat, and Longissimus thoracis (LT) muscle were collected for FA analysis and carcass and meat quality were measured. Compared with CT cattle, TT tended to have less (P = 0.06) C18:2-c9,t11 (rumenic acid) in plasma and subcutaneous fat and a greater proportion (P < 0.05) of C18:0 in subcutaneous, perirenal, and LT fat. Cattle with TT genotype also tended (P < 0.1) to have more total saturated and less unsaturated (USFA) and monounsaturated fats (MUFA) and had less (P = 0.04) linoleic acid in LT. Ultrasound fat thickness, REA, and average diameter of adipocytes in subcutaneous fat at 12 wk were not affected (P > 0.39) by genotype. Generally, carcass and meat quality were similar (P > 0.1) among diets, although adding FS tended to increase (P = 0.06) total USFA of subcutaneous fat including omega-3 FA (P < 0.001). For the high-fat diets evaluated, CT cattle would have more potential to produce beef with enhanced health benefits than would TT cattle.

Introduction

Leptin is an appetite and metabolism-related hormone that influences animal growth and fattening (Zhang et al., 1997). A single-nucleotide polymorphism (SNP) identified in exon 2 of the leptin gene encodes an amino acid substitution from arginine to cysteine (Buchanan et al., 2002). This SNP, commonly abbreviated LEP 25C (Kononoff et al., 2017), has been studied for impacts on performance and carcass characteristics of beef cattle. However, little information is available on the effects of this leptin SNP on adipocyte cellularity and fatty acid (FA) profiles including omega-3 FA. Omega-3 FAs such as α-linolenic acid (ALA) are essential for growth and early development and may play important roles in the prevention and treatment of coronary artery disease, hypertension, and other autoimmune disorders (Zárate et al., 2017). Flaxseed (FS) contains high concentrations of ALA and when added to finishing diets has increased ALA concentrations in plasma, fat, and muscle tissues of beef cattle (Kronberg et al., 2006; He et al., 2012a). High-oleate sunflower seeds (SS) are a rich source of linoleic acid (LA) which can be hydrogenated in the rumen to form conjugated linoleic acid (CLA; Gibb et al., 2004). Dried distillers grains with solubles (DDG) are a bioethanol byproduct and a source of concentrated protein, fat, and fiber widely used as ruminant feed (Klopfenstein et al., 2008) and have also been shown to increase ALA concentration in beef carcasses (He et al., 2012c). Dugan et al. (2010) found that supplementing beef diets with DDG decreased harmful trans-FA and increased CLA and vaccenic acid (VA), which may have anticancer effects (Xu and Qian, 2014). As leptin genotypes may possibly interact additively with dietary inclusion of oilseeds and DDG to influence carcass and meat quality traits, the hypothesis of the present study was that LEP 25C homozygous mutants (TT) would have increased subcutaneous fat accumulation and adipocyte cellularity compared with heterozygotes (CT) and differential responses to DDG and oilseeds in growth performance, meat quality, and FA profiles of various depots.

Materials and Methods

Experimental animals and determination of genotype

The study was conducted at the Lethbridge Research and Development Centre individual feeding barn and was reviewed and approved by the local Animal Care Committee, under the auspices of the Canadian Council of Animal Care (CCAC, 2009). Leptin genotype of cattle was determined based on tissue samples taken from the ear and analyses performed by Quantum Genetics Canada Inc., Saskatoon, SK, Canada as described by Kononoff et al. (2005). Leptin 25C genotypes include homozygous normal (CC), TT, and CT (McEvers et al., 2013). Of a group of 90 British-continental crossbred steers available for the study, LEP 25C genotypes were 12 CC, 35 CT, and 43 TT. Cattle used in this study were a subset of those in a companion study (He et al., 2012c, 2014) selected by balancing LEP 25C genotypes across DDG and oilseed diets.

Diets and animal feeding

The diets and their nutrient composition (dry matter [DM] basis) are presented in Table 1. Based on the available LEP 25C genotypes, 40 steers (20 CT and 20 TT; 459 ± 31 kg) were selected for the study. The experiment was designed as 2 × 2 × 2 factorial with three main factors: 1) genotype (CT or TT), 2) with or without inclusion of triticale DDG, and 3) with FS or SS supplementation. There were eight treatment groups and five steers per treatment. Steers received one of four diets for an average 15 wk before slaughter: 75% steam-rolled barley concentrate + 10% barley silage with 10% FS or SS and 46.5 % barley concentrate + 10% barley silage + 30% DDG, with 8.5% FS or SS. With the addition of a vitamin and mineral supplement, all diets fully met or exceeded the nutrient requirements of beef cattle as recommended by NRC (2000). Diets were formulated to be iso-caloric, equivalent in added fat, and <9% ether extract but were not iso-nitrogenous. Diets including DDG had increased concentrations of both CP and fiber compared with non-DDG (NDG) diets. Steers were housed individually, fed once daily at 8:30 a.m. and provided ad libitum access to their respective diets and water. For each pen, the amount of feed DM offered and orts were measured weekly to estimate the DM intake. Barley silage DM was monitored weekly and did not vary sufficiently (<2 %) to merit reformulation of the diet.

Table 1.

Experimental diets fed to CT and TT cattle (n = 5, per genotype and diet combination)

	NDG + FS	NDG + SS	DDG + FS	DDG + SS
Ingredient, % DM
Barley	75.0	75.0	46.5	46.5
Barley silage	10.0	10.0	10.0	10.0
FS	10.0	0.0	8.5	0.0
SS (high oleic)	0.0	10.0	0.0	8.5
Triticale DDG	0.0	0.0	30.0	30.0
Supplements1	5.0	5.0	5.0	5.0
Nutrients, % DM basis
DM, %	74.7	74.5	76.8	76.7
Protein, %	13.8	13.5	21.0	20.7
NEm2, Mcal/kg	2.0	2.1	2.1	2.2
NEg, Mcal/kg	1.3	1.4	1.4	1.5
Degradable carbohydrate, %	42.0	42.0	26.0	26.0
Ether extract, %	7.1	8.0	7.6	8.3
Calcium, %	0.6	0.6	0.7	0.6
FA, % total FAME
C14:0	0.07	0.06	0.06	0.05
C16:0	7.49	6.38	8.29	7.25
C18:0	2.40	3.37	2.25	3.07
C18:1-c9	15.36	54.12	16.54	48.53
C18:1-c11	0.73	0.77	0.77	0.80
C18:2 n-6	24.99	33.39	31.88	37.82
C20:1-c9	0.34	0.41	0.39	0.44
C18:3 n-3	48.61	1.50	39.83	2.04
SFA	9.97	9.80	10.60	10.37
USFA	90.03	90.20	89.40	89.63
MUFA	16.43	55.30	17.69	49.77
PUFA	73.61	34.89	71.71	39.86

¹The supplement provided 1 kg diet (in DM) with additional: 14.67 mg copper, 58.32 mg zinc, 26.73 mg manganese, 0.66 mg iodine, 0.23 mg cobalt, 0.29 mg selenium, 4,825 IU vitamin A, 478 IU vitamin D and 32 IU vitamin E.

²FAME, fatty acid methyl esters; NEg, net energy for growth; NEm, net energy for maintenance.

Tissue sampling and measurements

Steers were individually weighed prior to feeding at the start of the experiment and after 6 and 12 wk. Ultrasound measurements of subcutaneous fat and area of the Longissimus thoracis (LT) muscle and all blood and tissue samples were also collected prior to feeding and at 6 and 12 wk. Blood samples were collected from the jugular vein, and subcutaneous fat biopsies were collected surgically (He et al., 2012a). Adipocyte sizes in subcutaneous fat taken via biopsy were determined as described by He et al. (2010). Based on ultrasound fat thickness and bodyweight, the amount of fat trim was estimated (Realini et al., 2001). Blood samples were collected in the morning prior to feeding, using vacutainers (Becton Dickinson, Franklin Lakes, NJ, USA) containing Na₂-ethylenediaminetetraacetic acid. Blood plasma was then collected and 1 mL was stored at −40 °C pending extraction of lipids (He et al., 2011).

Carcass quality evaluations

Steers across dietary treatments were assigned to one of four slaughter dates at a commercial facility based on pen allocation, body weight, and initial ultrasound fat depth. Live weights were determined the day prior to transport to slaughter. Steers remained in lairage overnight within respective treatment groups and were given free access to water. At slaughter, they were stunned by captive bolt, exsanguinated, and dressed (Hernandez-Calva et al., 2011). Carcass quality traits, including carcass weight, dressing percentage, subcutaneous fat depth at the grade site, rib eye area (REA), marbling score, and quality grade, were recorded.

Meat quality traits on the carcass were further evaluated (Aldai et al., 2010; Hernandez-Calva et al., 2011). Within 45 min after slaughter, meat temperature and pH were measured posterior to the grade site on the left LT muscle. Carcasses were then pasteurized by exposure to steam at 105 °C for 6 s. Pasteurized carcasses were placed in a 2 °C cooler with a wind speed of 0.5 m/s and chilled for 24 h. Chilled carcass sides were then weighed. The left carcass sides were assessed for grade fat thickness, REA, and marbling score by a certified Canadian Food Inspection Agency grader (CFIA, 2001). Numeric marbling scores were also assessed according to the American Meat Science Association (1990) standards. Samples of LT muscle with attached subcutaneous fat and perineal fat samples were collected and frozen at −35 °C for subsequent FA analyses. The trimmed LT muscle samples were labeled, vacuum-packaged, and placed in the cooler at 2 °C for 6 d of aging.

Meat quality evaluations

Meat quality traits that were evaluated included shear force, color, drip loss, and nutrient concentration (Hernandez-Calva et al., 2011). After aging, the left LT was removed from the cooler and steaks (2.5 cm thickness) were taken from the posterior end. Pre-weighed steaks were grilled (Garland Grill ED30B; Condon Barr Food Equipment Ltd., Edmonton, AB) to reach an internal temperature of 35.5 °C. Steaks were turned and cooked to a final temperature of 71 °C (Hewlett Packard HP34970A Data Logger; Hewlett Packard Co., Boise ID) and then placed into polyethylene bags, sealed, and cooled by immersion in ice water. Steaks were then transferred to a cooler for 24 h, after which their weight was recorded for determination of cooking loss. Six 1.9-cm cores were used for peak shear force determination on the fiber grain (TA-XT Plus Texture Analyzer equipped with a Warner–Bratzler shear head) at a crosshead speed of 20 cm per min, with a 30 kg load cell, and using Texture Exponent 32 Software (Texture Technologies Corp., Hamilton, MA).

Color was measured on the steaks at 24 h and 144 h after slaughter. Three measurements such as color brightness (L*), color measurement of red-green axis (a*), and color measurement of yellow-blue axis (b*) were made with a chroma meter (CR-300, Konica Minolta, Ramsey NJ) across the surface of each steak after a 20-min bloom period. Color measurements were then converted to hue and chroma as described by Aldai et al. (2010). To determine drip loss, the steak was pre-weighed into a polystyrene tray with a Dri-Loc pad, over-wrapped with oxygen-permeable film, and stored for 5 d at 1 °C. The remaining portion of the LT was trimmed of all overlying connective tissue and ground three times using a Butcher Boy meat grinder with a 2-mm grind plate (Model TCA22, Lasar Manufacturing Co., Los Angeles, CA). Moisture was estimated by oven drying the remaining LT sample at 55 °C. For nutrient analyses, the dried LT sample was then ground (Grindomix Model GM200; Retsch Inc., Newton, PA). Crude fat (Method 960.39; Association of Official Analytical Chemists, 1995a) was analyzed via ether extraction (Foss Soxtec System Model 2050; Foss Analytical AB, Hoganas, Sweden) and nitrogen content (Method 992.15; Association of Official Analytical Chemists, 1995b) was measured by combustion analysis (CNS2000, Leco Corp., St. Joseph, MI).

Lipid extraction, methylation, and FA analysis

Feed samples were first freeze-dried prior to lipid extraction with ethyl ether using a Goldfisch apparatus (Laboratory Construction Co., Kansas City, MO, USA). Plasma lipids were extracted with isopropanol and hexane (He et al., 2011). Lipids in carcass tissues and muscles were extracted using previously described procedures (Radin, 1981; Kronberg et al., 2006, He et al., 2012b). For methylation, a combined acid/base methylation procedure using sodium methoxide (0.5 mol L⁻¹ in methanol) and boron trifluoride (14 % in methanol) was used (Lock and Garnsworthy, 2002). As an internal standard, nonadecanoic acid (C19:0) methyl ester (100 μL, 5.96 mg mL⁻¹ hexane Nu-Chek Prep, Inc., MN, USA) was added to lipid residues before methylation. Unless otherwise designated, chemicals were purchased from Sigma-Aldrich Inc. (Oakville, ON, CA). Fatty acid methyl esters were quantified (He et al., 2012a) using a gas chromatograph (Hewlett Packard GC System 6890, Mississauga, ON) equipped with a flame ionization detector and an SP-2560 fused-silica capillary column (75 m × 0.18 mm × 0.14 μm, Supelco Inc., Oakville, ON).

Statistical analyses

Average daily gain (ADG), feed intake (FI), feed conversion efficiency, subcutaneous fat thickness, FA composition of plasma and fat tissues, carcass quality, and meat quality data were analyzed using the MIXED procedure of SAS (Version 9.1, SAS Institute Inc., Cary, NC, 2004). The statistical model included genotype, DDG, oilseed (FS or SS), and all two-way interactions as fixed effects. Pen (individual) was the experimental unit. Means were separated by use of the Tukey test and a threshold of P < 0.05 was required for significant differences with P < 0.1 and > 0.05 considered a tendency.

Results

Animal growth performance

Including DDG reduced (P < 0.01) final weight, ADG, and feed conversion efficiency (G:F) compared with NDG but did not affect FI (Table 2). Genotype or two-way interactions including genotype and oilseed did not influence the performance.

Table 2.

Growth performance and ultrasound measurements for the 12-wk study

	Genotype		DDG		Oilseed			Treatment P-value
	CT	TT	No	Yes	FS	SS	SEM	G	DDG	OS
Body weight
Initial weight, kg	462.3	458.5	464.7	456.0	461.9	458.9	6.0	0.65	0.31	0.72
Final weight, kg	587.6	598.4	606.9	579.0	595.7	590.2	6.2	0.24	0.01	0.54
Performance
ADG kg/d	1.51	1.64	1.74	1.41	1.61	1.54	0.07	0.24	0.01	0.20
FI, kg/d	8.8	9.2	8.8	9.1	8.9	9.1	0.5	0.32	0.51	0.63
G:F kg/kg	0.16	0.17	0.19	0.15	0.17	0.16	0.01	0.33	<0.001	0.13
Ultrasound1
REA at 12 wk, cm2	78.6	78.3	80.2	76.8	78.6	78.3	2.1	0.93	0.28	0.93
Δ REA over 12 wk, cm2	38.4	41.5	41.3	38.6	39.5	40.4	2.5	0.40	0.44	0.82
BF at 12 wk, mm	14.2	13.9	14.8	13.2	14.6	13.4	1.4	0.78	0.11	0.21
Δ BF over 12 wk, mm	7.3	7.3	8.2	6.5	8.0	6.6	1.4	0.99	0.07	0.14

¹Ultrasound: REA, area of LT muscle.

Ultrasound measurements and adipocyte size

Genotype, oilseed, and DDG or their two-way interactions did not significantly (P > 0.1) affect ultrasound measurements of the subcutaneous fat thickness or REA during the 12-wk experimental period (Table 2). Average size and adipocyte hypertrophy after 12 wk were not affected (P = 0.1) by genotype or diet or by two-way interactions (Table 3). Similarly, genotype and two-way interactions between genotype, DDG, and oilseed did not affect (P > 0.1) adipocyte size distribution. Introduction of dietary DDG decreased (P = 0.02), whereas adding FS increased (P < 0.01) the proportion of adipocytes with diameters between 151 and 175 µm compared with SS.

Table 3.

Adipocyte enlargement and size distribution in the biopsy of subcutaneous fat at the end of the 12-wk study

	Genotype		DDG		Oilseed			Treatment P-value
Average diameter, µm	CT	TT	No	Yes	FS	SS	SEM	G	DDG	OS
Initial	127.4	128.7	126.2	129.9	130.7	125.4	7.0	0.82	0.44	0.28
At 12 wk	143.5	139.6	142.1	141.0	146.6	136.5	4.1	0.51	0.86	0.10
Δ over 12 wk	18.5	11.2	18.5	11.1	15.8	13.8	10.1	0.39	0.36	0.80
Cellular diameter distribution
26 to 75 µm	0.6	0.7	0.7	0.6	0.5	0.9	0.2	0.78	0.19	0.24
76 to 100 µm	8.1	9.9	9.1	8.8	7.2	10.7	1.8	0.48	0.94	0.18
101 to 125 µm	24.3	27.3	24.0	27.1	21.7	29.9	3.7	0.56	0.49	0.13
126 to 150 µm	26.4	26.7	24.8	28.2	24.6	28.5	1.6	0.88	0.65	0.11
151 to 175 µm	21.7	20.0	25.2	16.6	26.6	15.2	2.5	0.64	0.02	0.01
176 to 200 µm	13.4	10.7	13.2	10.9	14.5	9.6	2.3	0.42	0.48	0.15
201 to 250 µm	3.2	3.1	0.8	5.5	3.3	2.0	3.7	0.96	0.11	0.91

Carcass and meat quality

There were no significant differences (P < 0.05) among major carcass traits, including dressing percentage, cooler drip loss, grade fat, muscle score, ribeye area, marbling scores, and carcass quality grades among treatments (Table 4). The exceptions were fat class and dressing percent, which were increased (P < 0.05) by inclusion of flax as compared with SS. Similarly, most meat quality traits, including pH, temperature, and nutrient composition, were not affected (P > 0.10) by genotype, diet, or their two-way interactions (Table 5). The exception was inclusion of DDG, which increased (P = 0.05) the amount of drip loss and chroma at 144 h.

Table 4.

Carcass quality as affected by main effects of genotype after receiving experimental diets for 12 wk

	Genotype		DDG		Oilseed			Treatment P-value
	CT	TT	No	Yes	FS	SS	SEM	G	DDG	OS
Carcass, kg	354.9	346.2	358.6	342.4	358.7	342.3	10.0	0.38	0.09	0.09
Dressing %	60.0	60.1	59.8	60.4	60.6	59.7	0.30	0.75	0.09	0.02
Cooler loss, %	1.26	1.28	1.29	1.25	1.27	1.27	0.07	0.88	0.65	0.98
Grade fat, mm	19.8	19.7	20.2	19.3	21.3	18.1	1.2	0.94	0.59	0.08
Muscle score	2.5	2.3	2.3	2.5	2.5	2.3	0.3	0.74	0.64	0.64
Fat class	9.3	8.9	9.2	8.9	10.1	8.2	1.1	0.69	0.68	0.03
REA, cm2	81.0	80.6	81.6	79.8	81.8	79.6	2.1	0.89	0.53	0.50
Marbling score	527.3	540.8	531.1	537.1	531.2	536.9	10.5	0.38	0.69	0.71
AAA marbling1, %	88.0	80.2	83.5	84.5	79.5	88.5		0.99 (by χ 2 test)

¹AAA marbling %, A Canadian quality grade requiring small amounts of visible marbling.

Table 5.

Meat quality by main effects of genotype and dietary treatments measured on LT after 12 wk receiving experimental diets

	Genotype		DDG		Oilseed			Treatment P-value
	CT	TT	No	Yes	FS	SS	SEM	G	DDG	OS
Drip loss mg/g	26.7	29.1	25.7	30.1	27.5	28.4	1.5	0.26	0.05	0.68
Shear, kg	5.22	5.28	5.30	5.20	5.02	5.49	0.27	0.86	0.80	0.25
Cook loss, mg/g	173	177	173	177	173	177	8.0	0.72	0.69	0.69
Cook time, sec/g	3.55	3.70	3.47	3.78	3.46	3.79	0.37	0.78	0.55	0.52
Nutrient %
Moisture	70.9	71.0	70.9	71.0	71.0	70.9	1.1	0.97	0.93	0.96
Fat	6.7	6.8	6.7	6.8	6.8	6.7	1.3	0.97	0.81	0.89
Protein	21.7	21.7	21.8	21.6	21.6	21.7	0.1	0.75	0.21	0.64
pH, 45 min	6.81	6.86	6.86	6.80	6.82	6.85	0.1	0.26	0.14	0.51
pH, 24 h	5.58	5.60	5.59	5.60	5.60	5.58	0.02	0.11	0.78	0.30
Temperature °C
5 min	39.9	40.0	39.8	40.0	40.0	40.0	0.3	0.32	0.19	0.63
24 h	2.7	2.8	2.8	2.7	2.8	2.7	0.2	0.68	0.71	0.63
L* 1 24 h	38.5	38.9	38.5	38.9	38.7	38.7	1.8	0.71	0.66	0.93
L* 144 h	40.2	39.1	39.1	40.1	39.2	40.0	0.5	0.14	0.16	0.32
Chroma 24 h	23.8	23.6	23.3	24.1	24.1	23.3	0.3	0.58	0.17	0.21
Chroma 144 h	24.1	23.9	23.4	24.6	24.4	23.6	1.2	0.72	0.05	0.21
Hue 24 h	25.2	25.1	25.0	25.2	25.0	25.3	0.2	0.71	0.56	0.30
Hue 144h	24.8	24.7	24.7	24.8	24.7	24.8	0.3	0.87	0.76	0.94

¹L*, color brightness.

Plasma FA profiles

Type of oilseed affected (P < 0.001) proportions of total saturated fatty acids (SFA) and total unsaturated fatty acids (USFA) in blood plasma (Table 6). Compared with SS diets, cattle receiving FS had a greater proportion (P < 0.001) of USFA, less (P < 0.001) SFA, less (P < 0.01) monounsaturated fatty acids (MUFA), and more (P < 0.001) polyunsaturated fatty acids (PUFA) in plasma. In cattle with CT genotype, inclusion of DDG resulted in 47% less (P = 0.04) MUFA (Table 7). However, MUFA concentrations for the TT genotype were reduced by only 25% with DDG. Total omega-3 FA was affected (P < 0.001) by oilseed and inclusion of DDG. Inclusion of FS to diets increased (P < 0.001) the proportion of total omega-3 FA in plasma compared with that associated with the SS diet, whereas inclusion of DDG reduced (P < 0.001) total omega-3 FA. Cattle receiving the FS diet had more (P = 0.01) non-conjugated non-methylene interrupted dienes (NCD) compared with those receiving SS diet, although inclusion of DDG resulted in similar (P = 0.28) NCD between FS and SS diets. Proportions of total trans-FA and C18:1-trans were not affected (P > 0.1) by genotype or diet.

Table 6.

FA profile of plasma collected after 12 wk as affected by main effects of genotype and diet

	Genotype		DDG		Oilseed			Treatment P-value
FA1	CT	TT	No	Yes	FS	SS	SEM	G	DDG	OS
Total and major FA, % of FAME
SFA	30.8	30.8	29.8	31.9	26.5	35.2	0.9	0.99	0.12	<0.001
USFA	69.2	69.2	70.2	68.1	73.5	64.8	0.9	0.99	0.12	<0.001
MUFA	12.8	11.8	15.3	9.4	10.5	14.2	0.8	0.40	<0.001	0.01
PUFA	56.4	57.3	54.9	58.8	63.0	50.6	1.2	0.60	0.04	<0.001
Trans-FA	1.5	1.6	1.7	1.4	1.5	1.6	0.2	0.66	0.20	0.52
CLA & VA	0.30	0.28	0.23	0.35	0.25	0.33	0.05	0.76	0.09	0.23
Omega-3 FA	11.89	12.62	14.80	9.70	18.94	5.56	1.3	0.51	<0.001	<0.001
C18:1-trans	1.17	1.25	1.18	1.24	1.03	1.39	0.25	0.71	0.73	0.05
NCD	0.21	0.29	0.41	0.09	0.39	0.11	0.07	0.49	0.01	0.01
C16:0	8.1	8.0	8.1	8.0	7.2	8.9	0.5	0.94	0.72	0.001
C18:0	18.9	18.9	17.6	20.2	16.3	21.5	1.8	0.98	0.03	<0.001
C18:1-t10	0.54	0.63	0.63	0.53	0.52	0.64	0.06	0.29	0.22	0.15
C18:1-t11 (VA)	0.18	0.22	0.12	0.28	0.19	0.21	0.04	0.44	0.01	0.85
C18:1-c9	8.6	8.1	10.5	6.2	6.9	9.8	0.6	0.58	<0.001	0.01
C18:1-c15	0.7	0.5	0.8	0.3	1.0	0.1	0.1	0.36	0.01	<0.001
C18:2-c9,c12 (LA)	42.0	41.8	37.0	46.7	41.6	42.2	3.4	0.93	0.001	0.78
C18:2-c9,t11 (RA)	0.12	0.03	0.11	0.07	0.05	0.13	0.02	0.06	0.33	0.04
C18:2-t11,c15	0.08	0.13	0.18	0.03	0.19	0.02	0.04	0.38	0.01	0.01
C18:3n-3 (ALA)	8.79	9.49	11.49	6.78	15.5	2.8	1.4	0.53	<0.001	<0.001

¹FA: FAME, fatty acid methyl ester; Omega-3 FA, C18:3n-3 + C20:3n-3 + C20:5n-3 +C22:5n-3 + C22:6n-3; C:18:1-trans = coelution of several trans isomers.

Table 7.

Significant two-way interactions between genotype and diet for FA profiles across evaluated depots

		Genotype × diet
Depot1	FA	CT + NDG	CT + DDG	TT + NDG	TT + DDG	SEM	Interaction P-value
Plasma	MUFA	16.7d	8.8a	13.5c	10.1b	0.9	0.04
Plasma	C18:1c9	11.4c	5.6a	9.3c	6.9b	0.7	0.03
SF	ALA	0.40a	0.59b	0.42a	0.47a	0.03	0.07
LT	LA	1.34a	2.56c	1.37a	2.18b	0.08	0.02
LT	ALA	0.41a	0.57b	0.44a	0.48a	0.03	0.07
		CT + flax	CT + SS	TT + flax	TT + SS
Plasma	ALA	15.0b	2.6a	16.1c	2.4a	0.8	0.03

¹Depot: SF, subcutaneous fat.

^a–dMeans with different superscripts differ, P < 0.05.

High-oleate SS increased (P < 0.001) the proportion of C16:0 and C18:0 compared with FS, whereas genotype had no effect (P > 0.34) and DDG supplementation increased (P = 0.04) stearic acid in plasma. There was an interaction (P = 0.05) of genotype by DDG on C18:1-c9. Overall, inclusion of DDG decreased (P < 0.001) C18:1-c9; however, this effect was more marked (P = 0.05) for the CT as compared with TT genotype (Table 7). As well, proportion of C18:1-c9 was increased (P = 0.01) by inclusion of SS compared with FS. In complete opposition to C18:1-c9, C18:1-c15 increased (P < 0.01) in plasma from cattle receiving FS as compared with SS and was greater (P = 0.01) in cattle receiving NDG. An interaction between oilseed and DDG (P = 0.01) reduced C18:1-c15 in DDG + FS diets compared with NDG+FS but did not affect (P = 0.94) SS + DDG diets (Table 8). Inclusion of DDG increased (P < 0.001) LA. Cattle with CT genotype tended to have a larger proportion (P = 0.06) of rumenic acid (RA) in plasma compared with the TT genotype and this difference was significant in kidney fat (Table 9) but other fat depots assessed did not show similar genotype effects (Tables 10 and 11). Supplementing the CT genotype with flax reduced ALA (P < 0.05) in plasma compared with TT genotype fed flax, while ALA concentrations in both genotypes were similar (P > 0.1) when fed SS (Table 7). Supplementing with DDGs reduced proportion (P < 0.001) of ALA, but DDG + FS resulted in a lower (P < 0.05) concentration compared with NDG + FS (Table 8).

Table 8.

Significant two-way interactions between triticale distillers grains and oilseed for FA profiles across evaluated depots

		DDG × oilseed
Depot1	FA	NDG + FS	NDG + SS	DDG + FS	DDG + SS	SEM	Interaction P-value
Plasma	C18:1-c15	1.51c	0.17ab	0.55b	0.15a	0.16	0.01
Plasma	ALA	19.9c	2.6a	11.2b	2.4a	0.4	< 0.001
SF	NCD	2.84c	0.44a	1.63b	0.40a	0.17	0.01
SF	C18:2t11 c15	0.95c	0.07a	0.39b	0.06a	0.08	0.01
SF	ALA	0.66b	0.16a	0.88c	0.17a	0.04	0.01
LT	NCD	1.73c	0.26a	1.12b	0.26a	0.11	0.02
LT	C18:2t11 c15	0.41c	0.22a	0.36b	0.17a	0.06	0.02

¹Depot: SF, subcutaneous fat.

^a–cMeans with different superscripts differ, P < 0.05.

Table 9.

FA profiles in perirenal fat collected at slaughter after 12 wk receiving experimental diets as affected by main effects of genotype and diet

	Genotype		DDG		Oilseed			Treatment P-value
Fatty acid1	CT	TT	No	Yes	FS	SS	SEM	G	DDG	OS
Total and major FA, % of FAME
SFA	60.7	61.7	60.4	61.9	59.1	63.2	0.9	0.42	0.23	0.01
USFA	39.3	38.3	39.6	38.1	40.9	36.8	0.9	0.42	0.23	0.01
MUFA	35.7	34.7	36.3	34.1	35.8	34.6	0.8	0.41	0.07	0.30
PUFA	3.56	3.51	3.21	3.87	5.00	2.08	0.6	0.91	0.07	<0.001
Trans-FA	3.70	3.80	4.18	3.32	4.39	3.11	0.23	0.78	0.02	0.001
CLA & VA	0.87	0.86	0.80	0.92	0.89	0.83	0.12	0.89	0.15	0.45
Omega-3 FA	0.43	0.46	0.34	0.56	0.79	0.10	0.07	0.58	0.001	<0.001
C18:1-trans	2.81	2.92	3.08	2.64	3.00	2.72	0.16	0.61	0.07	0.24
NCD	0.69	0.71	0.89	0.51	1.20	0.20	0.09	0.89	0.01	<0.001
C16:0	28.2	28.3	27.7	28.7	28.0	28.4	0.6	0.92	0.23	0.59
C16:1-c9	1.42	1.26	1.42	1.27	1.35	1.33	0.08	0.17	0.20	0.84
C18:0	26.4	29.8	26.8	29.4	27.2	29.0	0.9	0.02	0.06	0.18
C18:1-t10	1.51	1.56	1.78	1.30	1.65	1.42	0.12	0.76	0.01	0.21
C18:1-t11 (VA)	0.68	0.71	0.62	0.77	0.72	0.66	0.11	0.77	0.05	0.41
C18:1-c9	29.4	28.5	29.6	28.3	29.1	28.8	0.8	0.46	0.24	0.73
C18:1-c15	0.66	0.62	0.66	0.62	1.20	0.08	0.11	0.81	0.77	<0.001
C18:2-c9,c12 (LA)	1.39	1.43	0.95	1.88	1.49	1.33	0.17	0.79	<0.001	0.25
C18:2-c9,t11 (RA)	0.19	0.15	0.18	0.16	0.17	0.18	0.02	0.04	0.16	0.65
C18:2-t11,c15	0.34	0.38	0.49	0.23	0.66	0.06	0.07	0.73	0.02	<0.001
C18:3n-3 (ALA)	0.42	0.45	0.32	0.54	0.77	0.09	0.07	0.59	0.001	<0.001

¹FA: FAME, fatty acid methyl ester; Omega-3 FA, C18:3n-3 + C20:3n-3 + C20:5n-3 +C22:5n-3 + C22:6n-3; C:18:1-trans = coelution of several trans isomers.

Table 10.

FA profiles in subcutaneous fat collected at slaughter after 12 wk receiving experimental diets as affected by main effects of genotype and diet

	Genotype		DDG		Oilseed			Treatment P-value
FA1	CT	TT	No	Yes	FS	SS	SEM	G	DDG	OS
Total and major FA, % of FAME
SFA	42.2	44.4	42.9	43.8	42.1	44.6	0.9	0.09	0.50	0.06
USFA	57.8	55.6	57.1	56.2	57.9	55.4	0.9	0.9	0.50	0.06
MUFA	53.3	51.4	52.8	52.0	51.9	52.9	0.8	0.11	0.46	0.37
PUFA	4.41	4.15	4.28	4.28	6.06	2.51	0.25	0.47	0.99	<0.001
Trans-FA	4.26	4.15	4.74	3.67	5.18	3.22	0.27	0.78	0.01	<0.001
CLA & VA	0.96	0.90	0.86	0.99	0.94	0.91	0.04	0.36	0.06	0.67
Omega-3 FA	0.54	0.48	0.44	0.57	0.82	0.19	0.03	0.21	0.01	<0.001
C18:1-trans	2.52	2.53	2.74	2.31	2.58	2.47	0.15	0.99	0.05	0.61
NCD	1.33	1.27	1.61	0.98	2.20	0.39	0.14	0.77	0.01	<0.001
C16:0	26.9	27.4	27.1	27.2	26.6	27.7	0.5	0.52	0.92	0.13
C16:1-c9	4.5	4.1	4.4	4.3	4.4	4.2	0.2	0.20	0.70	0.41
C18:0	10.1	11.5	10.3	11.3	10.2	11.3	0.4	0.03	0.09	0.06
C18:1-t10	1.4	1.4	1.7	1.2	1.5	1.3	0.1	0.92	0.01	0.30
C18:1-t11 (VA)	0.56	0.55	0.48	0.62	0.54	0.56	0.03	0.83	0.01	0.82
C18:1-c9	41.6	40.4	41.0	41.1	40.1	42.0	0.7	0.19	0.91	0.06
C18:1-c15	0.55	0.64	0.78	0.41	1.11	0.08	0.12	0.59	0.04	<0.001
C18:2-c9,c12 (LA)	1.37	1.26	0.86	1.77	1.36	1.26	0.15	0.39	<0.001	0.41
C18:2-c9,t11 (RA)	0.31	0.26	0.33	0.25	0.45	0.13	0.03	0.27	0.07	<0.001
C18:2-t11,c15	0.35	0.36	0.50	0.21	0.66	0.05	0.07	0.93	0.01	<0.001
C18:3n-3 (ALA)	0.50	0.45	0.41	0.54	0.78	0.17	0.03	0.22	0.01	<0.001

¹FA: FAME, fatty acid methyl ester; Omega-3 FA, C18:3n-3 + C20:3n-3 + C20:5n-3 +C22:5n-3 + C22:6n-3; C:18:1-trans = coelution of several trans isomers.

Table 11.

FA profile of LT muscle at slaughter after 12 wk receiving experimental diets as affected by main effects of genotype and diet

	Genotype		DDG		Oilseed			Treatment P-value
FA1	CT	TT	No	Yes	FS	SS	SEM	G	DDG	OS
Total and major FA, % of FAME
SFA	45.4	47.0	46.4	46.0	45.4	47.0	0.6	0.07	0.58	0.07
USFA	54.6	53.0	53.6	54.0	54.6	53.0	0.6	0.07	0.58	0.07
MUFA	50.0	48.7	49.4	49.2	48.5	50.1	0.5	0.09	0.78	0.03
PUFA	4.6	4.3	4.1	4.8	6.0	2.9	0.2	0.33	0.04	<0.001
Trans-FA	3.3	3.4	3.7	2.9	3.9	2.7	0.2	0.64	0.02	0.01
CLA & VA	0.76	0.77	0.75	0.79	0.71	0.72	0.05	0.14	0.08	0.94
Omega-3 FA	0.66	0.60	0.56	0.69	1.00	0.26	0.02	0.07	0.01	<0.001
C18:1-trans	2.14	2.30	2.45	1.98	2.25	2.19	0.16	0.50	0.05	0.78
NCD	0.82	0.83	0.98	0.68	1.41	0.24	0.09	0.95	0.03	<0.001
C16:0	27.4	27.4	27.6	27.1	27.1	27.7	0.4	0.97	0.41	0.33
C16:1-c9	3.71	3.43	3.70	3.44	3.69	3.45	0.37	0.32	0.27	0.31
C18:0	12.7	14.0	13.0	13.7	12.8	13.9	0.35	0.02	0.18	0.05
C18:1-t10	1.32	1.34	1.58	1.08	1.38	1.27	0.12	0.91	0.01	0.51
C18:1-t11 (VA)	0.38	0.51	0.37	0.51	0.44	0.44	0.05	0.06	0.04	0.86
C18:1-c9	40.3	39.4	39.4	40.4	38.7	41.0	0.5	0.24	0.23	0.01
C18:1-c15	0.62	0.62	0.76	0.48	1.17	0.08	0.12	0.98	0.10	<0.001
C18:2-c9,c12 (LA)	1.96	1.77	1.34	2.39	1.97	1.77	0.06	0.04	<0.001	0.03
C18:2-c9,t11 (RA)	0.26	0.25	0.26	0.25	0.26	0.26	0.01	0.63	0.63	0.88
C18:2-t11,c15	0.27	0.29	0.38	0.19	0.52	0.05	0.06	0.84	0.02	<0.001
C18:3n-3 (ALA)	0.49	0.46	0.42	0.53	0.81	0.14	0.05	0.37	0.01	<0.001
C20:4n-6	0.27	0.22	0.21	0.28	0.23	0.27	0.03	0.22	0.08	0.30
C20:5 (EPA)	0.04	0.04	0.04	0.04	0.06	0.03	0.01	0.49	0.83	0.001
C22:5 (DPA)	0.11	0.09	0.09	0.11	0.12	0.08	0.03	0.38	0.33	0.04
C22:6 (DHA)	0.007	0.008	0.006	0.009	0.007	0.008	0.002	0.89	0.11	0.80

¹FA: DHA, docosahexaenoic acid; DPA, docosapentaenoic acid; EPA eicosapentaenoic acid; FAME, fatty acid methyl ester; Omega-3 FA, C18:3n-3 + C20:3n-3 + C20:5n-3 +C22:5n-3 + C22:6n-3; C:18:1-trans = coelution of several trans isomers.

Subcutaneous FA profiles

Type of oilseed affected FA in subcutaneous fat, as inclusion of FS increased (P < 0.001) total PUFA, trans-FA, omega-3 FA, and NCD (Table 10). Similar to FS, inclusion of DDG also increased (P = 0.01) total omega-3 FA but DDG decreased (P = 0.01) NCD (P = 0.01), with largest decreases (P < 0.05) occurring when FS was also included in the diet (Table 8).

Proportion of C18:0 was affected (P = 0.03) by genotype and was greater (P < 0.05) in TT cattle compared with CT. Cattle with genotype CT had a greater proportion (P = 0.05) of ALA compared with TT, but only when consuming the DDG diet (Table 7). However, the most abundant FA, C18:1-c9, was not affected (P > 0.19) by diet or genotype. Adding DDG decreased (P = 0.01) C18:1-t10 and increased (P < 0.001) the proportion of LA. A major biohydrogenation intermediate of ALA, C18-c15, was increased (P < 0.001) by inclusion of FS. Inclusion of FS also significantly increased (P < 0.001) ALA and C18:2-t11,c15 (P = 0.01) compared with SS. However, DDG + FS decreased (P < 0.05) C18:2-t11,c15 in subcutaneous fat compared with NDG + FS, but further increased (P <0.05) ALA (Table 8).

Perirenal FA profiles

Similar to plasma and subcutaneous fat FA profiles, total SFA, USFA, PUFA, trans-FA, omega-3, and NCD in perirenal fat were not affected (P > 0.1) by genotype as a main effect or interaction (Table 9). No two-way interactions between genotype, DDG, and oilseed were significant for perirenal fat. However, for individual FA, C18:0 was greater (P = 0.02), whereas RA was reduced (P = 0.04) in TT genotype perirenal fat compared with CT. Similar to effects on subcutaneous fat, SS diets decreased (P < 0.05) total USFA, PUFA, trans-FA, omega-3 FA, and NCD. Inclusion of DDG also increased (P < 0.05) omega-3 FA but decreased (P < 0.05) NCD.

Evaluating individual FA, inclusion of DDG tended to increase (P = 0.06) C18:0 but decreased (P < 0.05) C18:1-t10 in perirenal fat, although C18:1-c9 and C16:0 were not affected (P > 0.23) by the treatments. Supplementing with FS resulted in elevated (P < 0.001) C18-c15 compared with SS. Inclusion of DDG increased (P < 0.01) LA and VA but decreased (P = 0.01) C18:1-t10. Similar to subcutaneous fat, inclusion of FS significantly increased (P < 0.001) ALA and C18:2-t11,c15 compared with SS. There was an interaction (P < 0.05) of DDG by oilseed affecting ALA and C18:2-t11,c15.

Longissimus thoracis FA profiles

Genotype tended to affect (0.05 < P < 0.1) proportion of total SFA, USFA, and MUFA in LT muscle (Table 11), with TT cattle having a larger proportion of SFA and CT having greater USFA and MUFA. FS increased (P < 0.001) total PUFA and total omega-3 FA (P < 0.001) but resulted in less (P = 0.03) MUFA. FS also increased (P < 0.01) total trans-FA and NCD compared with SS. Inclusion of DDG increased (P = 0.04) PUFA. Inclusion of DDG to FS diet decreased (P < 0.05) NCD compared to NDG + FS, whereas adding DDG to SS did not affect these FA (P = 0.97; Table 8).

For individual FA, proportion of C18:0 was greater (P = 0.02) in TT as compared with CC cattle (Table 11). Cattle with TT genotype also tended to (P = 0.06) have more VA compared with CT in LT. The most abundant SFA in LT, C16:0, was not affected (P > 0.05) by genotype or diet. However, FS supplementation decreased (P = 0.01) C18:1-c9 but increased (P < 0.001) C18:1-c15. LA was greater (P = 0.02) for CT as compared with TT genotype (Table 7). LA was a greater proportion of FA (P = 0.03) in FS compared with SS diets. Similarly, C18:2-t11,c15 was an increased proportion of FA (P < 0.001) in diets supplemented with FS. Inclusion of FS increased (P < 0.001) ALA whereas there was an interaction (P = 0.05) of genotype by DDG affecting ALA concentrations. In cattle with CT genotype, inclusion of DDG further increased (P = 0.05) ALA, whereas in TT cattle DDGs addition did not affect (P = 0.28) ALA concentrations (Table 7). Adding DDG decreased (P = 0.01) C18:1- t10 but increased (P = 0.04) VA. Inclusion of DDG increased (P < 0.05) LA in both genotypes, although the increase was largest (P = 0.002) for CT (Table 7). Overall, C18:2-t11,c15 was decreased (P = 0.02) by addition of DDG to diets but increased (P < 0.001) by adding FS. Combining DDG and FS further reduced (P < 0.05) C18:2-t11,c15 concentrations which were not changed by adding DDG to SS (Table 8).

Discussion

Genotype, growth performance, and carcass quality

From an early study of the LEP 25C SNP, Buchanan et al. (2002) found that the TT genotype was associated with a fatter carcass, whereas CC cattle had leaner carcasses. A recent study of 2,948 crossbred steers also found that the TT genotype had increased intramuscular fat, yield grade, DM intake, and hot carcass weight than CC cattle (Kononoff et al., 2017). The present study evaluated TT and CT which have also shown differences in carcass and performance traits in the previous studies. Kononoff et al. (2005) found that TT cattle had greater carcass yield grades compared with CT, while Nkrumah et al. (2005) found that TT genotype cattle had increased FI, subcutaneous fat thickness, and marbling score compared with both CC and CT.

Even though a number of studies of LEP 25C found that TT cattle had increased carcass fatness and FI, the population of cattle evaluated, diets, and slaughter weights have likely influenced results. Shin and Chung (2007) found that in Korean cattle, CC genotype had greater subcutaneous fat thickness and marbling score compared with TT. In a population of Irish crossbred cattle, Pannier et al. (2009) found no association with any of the leptin SNP and intramuscular fat. Woronuk et al. (2012) found that TT cattle had the lightest carcasses weights of all genotypes, while Kononoff et al. (2013) found no differences among LEP 25C genotypes for FI or growth performance. Also highlighting the variability of results, McEvers et al. (2013) found that CT cattle had a lower ADG and FI than both CC and TT animals. For the crossbred British × Continental steers used in the present study, genotype as a main effect or as part of a two-way interaction did not impact any measure of performance or carcass quality, similar to the studies by Pannier et al. (2009) and Kononoff et al. (2013). As impacts of triticale DDG, FS and SS on performance, and carcass quality were presented in a companion study (He et al., 2012c), these will not be discussed in depth to avoid replication.

Similar FI across genotypes in the present study may have limited variation in measures of carcass fatness, although McEvers et al. (2013) found increased marbling and a greater proportion of USDA Choice carcasses in TT as compared with CC cattle having the same FI. As bovine leptin is mainly secreted by adipocytes (Ji et al., 1998), the similar grade fat depth, carcass dressing percentage, and marbling across genotypes may have minimized differences in secretion of leptin by TT and CT. Leptin polymorphisms not only have reduced the efficiency of binding of leptin to its receptor but have also increased activity related to wild-type leptin (Reicher et al., 2012). Accordingly, many factors influence the prediction of metabolic outcomes based on the leptin genotype.

As inclusion of DDG increased dietary lipids compared with NDG diets, the amount of oilseeds in DDG diets was slightly reduced from 10% to 8.5% to maintain total dietary lipid below 9% of DM and to avoid affecting FI (NRC, 2001). Although fat content of all diets was relatively high (7.1% to 8.3%), inclusion of 15% FS to replace barley grain in a companion study did not affect the growth performance of beef cows compared with controls (He et al., 2012a) and oilseed type did not impact growth performance in the present study. Sunflower seeds including high-oleic SS and high-linoleic SS have replaced barley grain in diets of feedlot cattle (Gibb et al., 2004; Shah et al., 2006; Mir et al., 2008; He et al., 2014) with no adverse effects on growth performance or meat quality.

In contrast to neutral impacts of oilseed in previous studies, both ADG and G:F were reduced in the present study by inclusion of DDG, similar to a companion study (He et al. 2012c). Inclusion of DDGs into diets of feedlot cattle has had inconsistent effects on growth performance; although at rates of 20% or less, these have been minimal (Gibb et al., 2008). The relatively high fat content of diets likely had most impact on the digestibility of fiber-rich DDG, leading to the reductions in performance noted with DDG diets.

Adipocyte size

Size influences the metabolic function of adipocytes, with large cells having 3-fold greater expression of leptin than small cells (Farnier et al., 2003) and other gene transcripts having up to 22-fold greater expression in large adipocytes (Jernås et al., 2006). As differences in number and size of adipocytes may confound comparisons of gene products such as leptin (Dodson et al., 2014), size and growth of adipocytes were measured in subcutaneous fat in the present study. Although adipocyte morphology is in part determined genetically (Tandon et al., 2018), CT and TT genotypes did not differ in average adipocyte size, adipocyte hypertrophy over 12 wk, or distribution of various sizes of adipocytes. Only minor effects on adipocyte size were shown, with DDG supplementation reducing and FS supplementation increasing the proportion of 151 to 175 µm adipocytes. Size of adipocytes differs among fat depots (Dodson et al., 2014), and fat depots of cattle differ in expression of leptin and other factors influencing adipogenesis (Romao et al., 2012). Consequently, adipose morphology in depots other than subcutaneous fat may have differed between CT and TT genotypes. However, as CT and TT also had similar carcass weight, dressing percentage, marbling score, total fat in LT, and subcutaneous fat depth measured by ultrasound, differences in adipocyte size or the number between genotypes are unlikely to have influenced the results of the present study.

Meat quality

Although Li et al. (2013) showed that the LT muscle of TT cattle had lower chroma and oxymyoglobin content as compared with CC or CT cattle, these authors did not observe other leptin genotype differences in meat quality. Intramuscular fat content has influenced meat color attributes (Fiems et al., 2000) and as marbling scores were similar between genotypes in the present study, meat color would be unlikely to differ. Similar to the present study, Li et al (2013) found no impact of leptin SNP on the water-holding capacity of meat. The only differences in meat quality noted in the present study were related to DDG supplementation. Drip loss was increased as was chroma at 144 h in LT from cattle receiving dietary DDG. Increased chroma and hue have been previously reported in LT muscle from cattle receiving corn- or wheat-based DDG (Aldai et al., 2010), although these differences were thought to be due to a greater ultimate pH in the carcasses of DDG-fed cattle. As carcass pH at 24 h did not differ among treatments in the present study, other factors would be responsible for the meat color differences observed. Adding 40% to 50% corn DDG to diets has reduced meat color stability (Roeber et al., 2005), but chroma and hue remained similar at 24 and 144 h in the present study and additions of DDG were restricted to 30% of DM. Although factors influencing drip loss and chroma at 144 h are unknown, He et al. (2014) noted similar changes to meat quality in a companion study when also feeding triticale DDG and that SS and FS dietary oilseeds did not impact carcass or meat quality.

Fatty acid analyses

The present study indicated that the LEP 25C genotype or interactions of genotype by DDG or oilseed could affect FA profiles of plasma, subcutaneous fat, perirenal fat, and lipids from LT muscle. However, genotype effects were rarely consistent across all fat depots measured. Cattle with TT genotype had reduced plasma and perirenal RA compared with CT cattle and increased C18:0 in subcutaneous, perirenal, and LT fat. Only in the LT were there trends (P < 0.1) for TT to have a greater concentration of SFA and less USFA, MUFA, and omega-3 FA. For other individual FA, CT had more LA only in LT fat. Of all tissues examined, FA profile of the LT would reflect the intramuscular “taste” fat, which adds value to a carcass (Aldai et al., 2007) and not the waste fat, which is trimmed or removed prior to retail fabrication. The intramuscular fat of the LT along with a portion of subcutaneous fat would be the only FA evaluated that would be included in meat products (da Silva Martins et al., 2018). Although the health effects of LA are in some cases contradictory (Jandacek, 2017), reducing saturated fat has long been the foundation of dietary recommendations for reducing cardiovascular disease (Liu et al., 2017). Accordingly, FA profiles of CT meat products would likely have more beneficial human health effects than would those of TT.

Excluding impacts of diet or genotype, FA profiles have long been known to differ among depots or even in different locations within a depot (Callow, 1962). It is only more recently that some of the mechanisms responsible for differential adipogenesis among depots have been investigated, including altered microRNA expression (Romao et al., 2012) and expression of stearoyl CoA desaturase-1, a key lipogenic gene (Petrus et al., 2017). For cattle, the fat depot evaluated has had a greater influence on the FA profile than does diet (Romao et al., 2013). Leptin has been found to inhibit expression of FA synthase and sterol regulatory element-binding protein (SREBP1) in mice (Soukas et al., 2000), but bovine and murine lipogeneses differ substantially (Joseph et al., 2010). Consequently, mechanisms whereby LEP 25C altered FA profiles of various depots require further investigation.

Compared with the high-oleic SS, FS provided a FA profile with much higher PUFA (mainly ALA), which resulted in greater plasma PUFA including total omega-3 FA. From previous companion studies, ALA concentration in diet influenced that in plasma of beef cows at 3 (He et al., 2011) and 24 h (He et al., 2012a) after feeding. In the present study, FS also increased ALA and total omega-3 FA in subcutaneous fat and LT muscle, similar to the previous studies (Kronberg et al., 2006; Juárez et al., 2011; Nassu et al., 2011). In a companion study, He et al. (2012c) found that compared with SS, inclusion of FS resulted in much greater ALA and total omega-3 FA which was also accompanied by increased ALA biohydrogenation intermediates with no difference in CLA. Generally, cattle diets containing greater concentrations of ALA may generate relatively greater concentrations of CLA during rumen fermentation, and inclusion of high linoleic SS but not high oleic SS to diets has increased beef CLA concentration (Shah et al., 2006; Mir et al., 2008).

Inclusion of wheat DDG to feedlot diets has affected beef FA profile, that is, increased ALA (He et al., 2012b), CLA, and the ratio of VA to 18:1-t10 (Aldai et al., 2010; Dugan et al., 2010). In the present study, inclusion of triticale DDG to FS but not SS diets increased ALA and total omega-3 FA in subcutaneous fat and LT muscle, whereas it decreased the major bio-hydrogenation intermediates of ALA, the NCD. Different from the effect on ALA in fat tissues, treatments with DDG diets had less ALA and total omega-3 FA in the plasma compared to those with NDGS diets. The reduced ALA in plasma by inclusion of 30% DDG to substitute barley (28.5%) and a small proportion (1.5%) of FS could be caused either by slightly lower ALA intake in DDG diets or more likely by more effective utilization of plasma ALA for incorporation into adipocytes compared with NDG cattle. More interestingly, the present study demonstrated that the effect of DDG on beef LA, ALA, and total omega-3 was affected by leptin genotype, which had a larger effect in CT compared with TT cattle.

Flax is known as a rich source of LA (Liu et al., 2017) and supplementing DDG also increased LA in all depots measured. Only in the LT depot did feeding DDG increase LA to a greater extent for CT as compared with TT cattle. However, independent of diet, CT cattle also had increased LA in LT fat. Leptin has influenced both SREBP1 and the receptor PPARγ which regulate expression of lipogenesis enzymes (Kim and Spiegelman, 1996). Accordingly, differential leptin activity among genotypes may have possibly led to increased LA in the LT of CT cattle. The role of leptin has been reported to differ among depots (Harris, 2014), which may explain why increased deposition of LA in CT cattle was only apparent in the LT.

It was not clear why including DDG + FS could increase ALA concentration in beef fat compared with inclusion of FS only. As a result of inclusion of DDG, dietary CP increased from 14% to 21%, whereas degradable carbohydrate decreased from 42% to 26%. This could have a significant effect on rumen fermentation and the biohydrogenation of dietary PUFA (Dugan et al., 2011) and caused changes to amounts and ratios of rumen volatile fatty acids (VFA). In bovine adipocytes, VFA are the major substrates for de novo FA synthesis (da Silva Martins et al., 2018). Compared with propionate, acetate contributes more to fat synthesis (Dugan et al., 2011) and, therefore, the possible decrease in acetate:propionate by inclusion of DDGs (Li et al., 2011) could result in less accumulation of those nonessential fatty acids from de novo synthesis and an increased incorporation rate of PUFA including ALA to adipocytes from plasma.

Conclusions

The present study was conducted to determine if the LEP 25C genotypes CT and TT had differential responses to dietary DDG and oilseeds which would affect growth performance, meat quality, and FA profiles of various depots. A better understanding of LEP 25C genotype impacts was needed as previous studies had conflicting results regarding growth performance and knowledge of LEP 25C impacts on FA and adipocyte cellularity was lacking. By evaluating FA profiles and carcass fat deposition, this study identified the potential for the LEP 25C genotypes evaluated to meet consumer demand for meat with enhanced concentrations of health-promoting FA.

As growth performance and carcass characteristics including adipocyte size, marbling, and rate of fat deposition were not affected by genotype or interactions of diet by genotype, the hypothesis that these would differ among genotypes and that TT carcasses would have additional fat was not supported. In contrast, the hypothesis that LEP 25C genotypes would differ in FA profiles shows merit. However, the impacts of LEP 25C genotypes on FA profiles were not uniform and were dramatically influenced by the depot evaluated. Differential adipogenesis according to LEP 25C genotype requires future study, although average adipocyte size, adipocyte hypertrophy over 12 wk, and distribution of various sizes of adipocytes did not differ among genotypes. Results of the current study demonstrate that different genotype and diet combinations could be used to produce desired outcomes. If carcass FA profiles for improved human health were the goal, CT cattle fed flax and/or DDG should be selected, although adding DDG to high-fat diets may result in reduced growth performance.

Glossary

Abbreviations

ADG

average daily gain

ALA

α-linolenic acid

BF

subcutaneous fat depth

CC

homozygous normal leptin genotype

CLA

conjugated linoleic acid

CT

heterozygous leptin genotype

DDG

dried distiller’s grains with solubles

DM

dry matter

FA

fatty acid

FI

feed intake

FS

flaxseed

G

genotype

G:F

feed conversion efficiency

LA

linoleic acid

LEP 25C

SNP in exon 2 of leptin gene with transition of cytosine to thymine

LT

Longissimus thoracis muscle

MUFA

monounsaturated fatty acids

NCD

non-conjugated non-methylene interrupted dienes

NDG

diets without DDG

OS

oil seed

PUFA

polyunsaturated fatty acids

RA

rumenic acid

REA

rib eye area

SFA

saturated fatty acids

SNP

single nucleotide polymorphism

SS

high-oleate sunflower seeds

TT

homozygous mutant leptin genotype

USFA

unsaturated fatty acids

VA

vaccenic acid

VFA

volatile fatty acids

Conflict of interest statement

The authors with the exception of L.M. declare no conflicts of interest. L.M. provides fee for service detection of leptin genotypes but in no way influenced the content of the manuscript.