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Liver metabolism traits in two rabbit lines divergently selected
for intramuscular fat
M. Martínez-Álvaro
1
, Y. Paucar
1
, K. Satué
2
, A. Blasco
1
and P. Hernández
1†
1
Institute for Animal Science and Technology, Universitat Politècnica de València, 46022 Valencia, Spain;
2
Department of Animal Medicine and Surgery, Universidad
Cardenal Herrera, 46113 Valencia, Spain
(Received 10 May 2017; Accepted 20 September 2017)
Intramuscular fat (IMF) has a large effect in the sensory properties of meat because it affects tenderness, juiciness and flavour.
A divergent selection experiment for IMF in
longissimus dorsi
(LD) muscle was performed in rabbits. Since liver is the major site of
lipogenesis in rabbits, the objective of this work is to study the liver metabolism in the lines of the divergent selection experiment.
Intramuscular fat content, perirenal fat weight, liver weight, liver lipogenic activities and plasma metabolites related to liver
metabolism were measured in the eighth generation of selection. Direct response on IMF was 0.34 g/100g of LD, which
represented 2.7 SD of the trait, and selection showed a positive correlated response in the perirenal fat weight. High-IMF line
showed greater liver size and greater liver lipogenic activities of enzymes glucose-6-phosphate dehydrogenase and malic enzyme.
We did not find differences between lines for fatty acid synthase lipogenic activity. With regard to plasma metabolites, low-IMF
line showed greater plasma concentration of triglycerides, cholesterol, bilirubin and alkaline phosphatase than high-IMF line,
whereas high-IMF line showed greater albumin and alanine transaminase concentrations than low-IMF line. We did not observe
differences between lines for glucose, total protein and plasma concentrations. Phenotypic correlations between fat (IMF and
perirenal fat weight) and liver traits showed that liver lipogenesis affects fat deposition in both, muscle and carcass. However,
the mechanisms whereby liver lipogenesis affected IMF content remain to be clarified.
Keywords: intramuscular fat, liver, metabolism, genetic selection, rabbits
Implications
Intramuscular fat (IMF) is a main factor in meat quality
because it affects sensory properties of meat. Genetic selection
for IMF in rabbits modifies liver size and lipogenic activity,
particularly the activity of the enzyme glucose-6-phosphate
dehydrogenase (G6PDH). Our study shows that liver plays
a main role in the genetics of IMF deposition in rabbits.
Introduction
Intramuscular fat has a large effect in the sensory properties of
meat. A high IMF content has been associated with tender,
juicy and flavourful meat in sheep, cattle and pig (Wood
et al
.,
2008). In rabbits, Hernández
et al
. (2000) reported a positive
relationship between IMF and juiciness. Intramuscular fat
can be easily modified by genetic selection, although there
are only three selection experiments for IMF published
(Schwab
et al
., 2009 in pigs, Sapp
et al
., 2002 in cattle and
Zhao
et al
., 2007 in chickens). In the Universitat Politècnica de
València we are performing a divergent selection experiment
for IMF in rabbits (Martínez-Álvaro
et al
., 2016).
Liver tissue is a major site of lipogenesis in some species
such as chickens (O’Hea and Leveille, 1969), rats (Ballard
et al
., 1969) and growing rabbits (Gondret
et al
., 1997). In
these species, IMF deposition may depend not only on the
metabolism of intramuscular adipocytes, but also on meta-
bolic activity of liver. Differences on lipogenic activities in
liver have been related to differences in IMF in chickens
(Cui
et al
., 2012), and to differences in fat depots in rats
(Smith and Kaplan, 1980; Turkenkopf
et al
., 1980) and pigs
(Muñoz
et al
., 2013). Our hypothesis is that the different IMF
deposition in the divergent rabbit lines of our experiment
would be related to different lipogenic activities in liver. To
test this hypothesis, we propose to measure in both lines
lipogenic enzyme activities in liver, and plasma metabolites
that are related to lipogenesis. The advantage of comparing
divergent lines selected for IMF is that they only differ in IMF
and correlated traits; therefore, differences between lines can
be only attributed to differences in IMF metabolism.
†
E-mail: phernan@dca.upv.es
Animal
, page 1 of 7 © The Animal Consortium 2017
doi:10.1017/S1751731117002695
animal
1
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Material and methods
Animals
A divergent selection experiment for IMF in muscle
longissimus dorsi
(LD) was performed in rabbits. A male and
a female from the first parity of each doe were slaughtered at
9 weeks of age and evaluated for IMF, and the average
between these two values was calculated. Then, all dams
were ranked according to this average, and selection for high
or low IMF was performed on rabbits from the second parity.
All females of the ~20% best dams were selected for next
generation. As each sire was mated with five dams, only one
male of its best dam was selected. This selection within
male family was performed in order to reduce inbreeding.
Normally, the first parity was used to collect the IMF data and
the second parity to select the rabbits for next generation,
although exceptionally some IMF measurements were made
on the second or third parity. Lines selected for high IMF and
low IMF were reared contemporary at the farm of the
Universitat Politècnica de València. The housing had a con-
stant photoperiod of 16:8h and controlled ventilation. Litters
were homogenized by performing adoptions at birth up to
9 kits per litter. From weaning to slaughter, rabbits were
reared collectively and fed
ad libitum
. More details of this
experiment can be found in Martínez-Álvaro
et al
. (2016).
This study was performed with 175 rabbits from the eighth
generation of this selection experiment, 83 from the high-
IMF line and 92 from the low-IMF line. Body weight was
recorded at 9 weeks of age. Then, all rabbits were fasted at
least 19 h before slaughtering by electrical stunning and
exsanguination. Carcasses were prepared according to the
norms of the World Rabbit Science Association (Blasco and
Ouhayoun, 1996). Carcasses were chilled for 24 h at 4°C and
the weight of the chilled carcass was recorded. Perirenal fat
depot was excised from the carcass and weighed. Muscle LD
was excised, minced, freeze-dried and scanned with NIRS
(model 5000; FOSS NIRSystems Inc., Hilleroed, Denmark).
Intramuscular fat was determined in g/100 g of muscle
applying the calibration equations previously developed by
Zomeño
et al
. (2011). The calibration reported for IMF had a
high precision and accuracy, according to the statistics
R
2
(0.98) and residual predictive deviation (7.57).
A subsample of 63 rabbits (30 from the high-IMF and 33
from the low-IMF line) was taken to study the liver lipogenic
activity and plasma metabolites. Animals were slaughtered as
described before. Blood samples were collected at slaughter
from the jugular vein in 1 ml lyophilized lithium heparin
(0.04 mg/ml) tubes (TapVal Aquisel, Barcelona, Spain) and
plasma was prepared by centrifugation at 3000r.p.m. for
10 min and then stored at −80°C. Liver was dissected from the
carcass and weighed immediately after slaughter. A liver
sample was frozen in liquid nitrogen, vacuum packed and
stored at −80°C for lipogenic enzyme assays.
All experimental procedures involving animals were
approved by the Universitat Politècnica de València Research
Ethics Committee, according to council directive 2010/63/EU
(European Commission Directive, 2010).
Lipogenic activities measurements
Activity of enzymes G6PDH (EC 1.1.1.49), malic enzyme (ME1;
EC 1.1.1.40) and fatty acid synthase (FASN; EC 2.3.1.85) were
measured. For ME1 and G6PDH measurements, 1 g of liver was
homogenized in 5 ml of ice-cold 0.25 M sucrose solution,
whereas for FASN measurement 0.5 g of liver was homo-
genized in 2.5 ml of ice-cold 0.25M sucrose solution containing
1 mM dithiothreitol and 1 mM ethylenediaminetetraacetic acid.
Homogenates were centrifuged at 12 000 gfor1hat4°Cand
supernatants were filtered through glass wool and collected for
enzyme assays. Lipogenic activities were assessed at 37°C
using a spectrophotometric analyzer Fluostar Galaxy (BMG Lab
Technologies, Offenburg, Germany) at 340 nm, according to
the method described by Zomeño
et al
. (2010) with some
modifications. Enzyme activities wereexpressedinnmolsof
NADP produced (G6PDH and ME1) or oxidized (FASN) per
minute and g of fresh tissue. Soluble protein was determined
in liver supernatant using the bicinchoninic acid Protein
Assay Kit provided by Thermo Fisher Scientific(Rockford,IL,
USA), and enzyme activities were also expressed in a
soluble-protein basis.
Plasma metabolites measurements
Fasting plasma concentrations (mg/dl) of glucose, total
cholesterol and triglycerides were determined by enzymatic
colorimetric methods. Glucose was determined by the
Trinder glucose oxidase method, triglycerides were measured
by the glycerol phosphate dehydrogenase –peroxidase
method and total cholesterol was measured by the choles-
terol oxidase –peroxidase method. Concentrations of
bilirubin (mg/dl), albumin (g/dl) and total protein (g/dl) were
determined by dimethyl sulfoxide, Bromocresol Green and
Biuret colorimetric methods, respectively. Finally, plasma
concentrations (UI/l) of enzymes aspartate transaminase
(AST; EC 2.6.1.1), alanine transaminase (ALT; EC 2.6.1.2) and
alkaline phosphatase (ALP; EC 3.1.3.1) were measured by
photometric methods. All the methods employed are
described in Kaplan
et al
. (2009). All the methodologies were
integrated in an automatic chemistry analyser model Spin
200E (Spinreact, Girona, Spain).
Statistical analysis
Descriptive statistics were estimated after correcting data by
the fixed effects of line and sex. Month-season and parity
order fixed effects were additionally included for IMF, BW,
chilled carcass and perirenal fat weights analysis. Direct
and correlated responses to selection were estimated as
the differences between high-IMF and low-IMF lines. All the
differences were estimated with a model including the
fixed effects of line, sex, month-season and parity order
(as described before) and common litter random effect.
Phenotypic correlations of IMF and perirenal fat weight with
liver weight, liver lipogenic activities and plasma metabolites
were estimated after correcting data for line and sex.
Bayesian inference was used (Blasco, 2017). Common
litter effect and residuals of the models were assumed to be
independently normally distributed. Bounded flat priors were
Martínez-Álvaro, Paucar, Satué, Blasco and Hernández
2
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assumed for all fixed effects and variances. Marginal
posterior distributions were estimated using Gibbs sampling.
Descriptive statistics and differences between lines were
performed with programme ‘Rabbit’, developed by the
Institute for Animal Science and Technology (Valencia,
Spain). After some exploratory analyses, results were based
on Monte Carlo Markov chains runs consisting of 60 000
iterations, with a burn-in period of 10 000, and only one of
every 10 samples were saved for inferences. Phenotypic
correlations were computed with the software TM (Legarra
et al
., 2008). In this case, after some exploratory analyses
results were based on Monte Carlo Markov chains runs
consisting of 1 000 000 iterations, with a burn-in period of
200 000, and only one of every 100 samples were saved for
inferences. Convergence was tested using the Z criterion of
Geweke and Monte Carlo sampling errors were computed
using time-series procedures.
The parameters obtained from the marginal posterior
distributions of the differences between lines and phenotypic
correlations were: the median, the highest posterior
density region at 95% (HPD
95%
) and the probability of the
difference or correlation being greater than 0 when the
median is positive or lower than 0 when the median is
negative (
P
0
). Additionally, we considered one-third of the
SD of a trait as a relevant value (
r
) and we calculated the
probability of relevance (probability of the difference
between lines being greater than
r
when the median is
positive or lower than
r
when the median is negative) (
P
r
). A
more detailed description of these features can be found in
Blasco (2017).
Results
Response to selection and correlated responses in
carcass traits
Table 1 shows descriptive statistics and differences between
lines for IMF and carcass traits. Direct response to selection
estimated as the difference between lines in the eighth
generation was 0.34 g/100 g of LD (
P
r
=1.00) with a HPD
95%
from 0.30 to 0.39. Expressed in units of SD, direct response
was 2.7 SD of the trait. Selection for IMF showed a positive
correlated response in the carcass adiposity. High-IMF
line showed greater perirenal fat weight (
P
0
=1.00) than
low-IMF line, and the difference between lines was relevant
(
P
r
=1.00). We did not find differences between lines in BW
and chilled carcass weight.
Liver weight and lipogenic activities
Table 2 shows descriptive statistics and differences between
lines for liver weight and liver lipogenic activities. The
greatest lipogenic activity in liver was G6PDH. High-IMF line
showed greater liver weight than low-IMF line (
P
0
=0.99)
and the probability of the difference between lines being
relevant was
P
r
=0.87. Besides, high-IMF line showed
greater G6PDH (
P
0
=1.00) and ME1 activities (
P
0
=0.92)
than low-IMF line. The only relevant difference between lines
was for G6PDH activity (
P
r
=1.00), showing a difference of
1182 nmol/min and g, or 1.51 SD of the trait. We did not find
differences between lines for FASN activity. Results were
similar when activities were expressed in a soluble-protein
basis (data not shown).
Plasma metabolites related to liver
Table 3 reports descriptive statistics and differences between
IMF rabbit lines for plasma metabolites related to liver. Low-
IMF line showed greater plasma concentration of triglyce-
rides, cholesterol, bilirubin and ALP than high-IMF line and
all the differences between lines were relevant, except for
cholesterol concentration, in which
P
r
was very low. High-
IMF line showed greater albumin and ALT concentrations
(
P
0
=1.00), and differences between lines were relevant. We
did not observe differences between lines for glucose, total
protein and AST plasma concentrations.
Relationships between fat and liver traits
Table 4 shows phenotypic correlations between fat traits
(IMF and perirenal fat weight) and liver traits (liver weight,
lipogenic activities and plasma metabolites). Intramuscular
fat was positively correlated with liver weight (P
0
=0.98)
and with G6PDH (P
0
=0.97) and FASN (P
0
=1.00) activities,
correlations ranging from 0.28 to 0.38. We do not have
enough evidence to state the sign of the correlation between
IMF and ME1 activity. Perirenal fat weight was positively
correlated with ME1 activity (0.34, P
0
=1.00). The correla-
tions between perirenal fat weight and G6PDH and FASN
activities and between perirenal fat and liver weights were
Table 1
Descriptive statistics and differences between high- and low-intramuscular fat(IMF) rabbit lines in IMF andcarcass traits (g)
Trait Mean SD D
1
HPD
95%2
P
03
r
4
P
r5
IMF 0.99 0.13 0.34 0.30, 0.39 1.00 4.36 1.00
BW 1750 112 7.50 −33.2, 47.9 0.64 2.13 0.07
Chilled carcass weight 974 80.3 12.5 −22.2, 47.9 0.75 2.75 0.20
Perirenal fat weight 7.77 2.36 3.19 2.35, 4.05 1.00 10.1 1.00
1
Median of the marginal posterior distribution of the difference between high-IMF and low-IMF lines.
2
Highest posterior density region at 95% of probability.
3
Probability of the difference being greater than 0 when D >0 or lower than 0 when D <0.
4
Relevant value, proposed as one-third of the SD of the trait.
5
Probability of relevance (probability of the difference being greater than
r
when D >0 or lower than
r
when D <0).
Metabolism of intramuscular fat
3
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also positive, but with lower evidence (
P
0
between 0.88 and
0.89) and showing lower values (from 0.16 to 0.17).
Albumin concentration in plasma was positively correlated
with IMF (0.27) and with perirenal fat weight (0.35) (Table 4).
Total protein plasma concentration had a low positive corre-
lation with IMF (0.21,
P
0
=0.94). Phenotypic correlations
between IMF and perirenal fat weight and the other plasma
metabolites measured were weak (data not shown).
Discussion
Divergent selection for IMF in rabbits was successful, as
previously observed in Martínez-Álvaro
et al
. (2016). The
genetic progress was approximately one-third of the SD of
the trait per generation. Selection for IMF showed a positive
and relevant correlated response in perirenal fat weight,
which is the main carcass fat depot in rabbits (Hernández
et al
., 2006). Other selection experiments for IMF also found
a positive correlated response in the carcass adiposity
(Schwab
et al
., 2009 in pigs and Zhao
et al
., 2007 in chick-
ens), and the positive genetic correlation between intra-
muscular and carcass fat is widely documented (Martínez-
Álvaro
et al
., 2016 in rabbits and Ciobanu
et al
., 2011 in a
pig review). High-IMF line showed greater liver size than
low-IMF line, which should be related to its greater fat
deposition, since liver is the tissue with the greatest lipogenic
activity in growing rabbits (Gondret
et al
., 1997).
Divergent selection for IMF allows studying the lipid
metabolism strictly underlying IMF deposition, since the
selected lines have the same genetic background and
only differ in genes involved in IMF and correlated traits.
Table 2
Descriptive statistics and differences between high- and low-intramuscular fat (IMF) rabbit lines in liver weight and liver lipogenic
1
activities
Trait Mean SD D
2
HPD
95%3
P
04
r
5
P
r6
Liver weight (g) 42.8 3.71 2.39 0.47, 4.50 0.99 2.88 0.87
G6PDH 4383 817 1182 698, 1660 1.00 272 1.00
ME1 416 102 44.8 −17.3, 108 0.92 33.8 0.64
FASN 686 83.0 9.60 −38.2, 56.9 0.65 27.7 0.22
1
Activities of the lipogenic enzymes glucose-6-phosphate dehydrogenase (G6PDH), malic enzyme (ME1) and fatty acid synthase (FASN) are expressed in nmol/min
and g of tissue.
2
Median of the marginal posterior distribution of the difference between high-IMF and low-IMF lines.
3
Highest posterior density region at 95% of probability.
4
Probability of the difference being greater than 0 when D >0 or lower than 0 when D <0.
5
Relevant value, proposed as one-third of the SD of the trait.
6
Probability of relevance (probability of the difference being greater than
r
when D >0 or lower than
r
when D <0).
Table 3
Descriptive statistics and differences between high- and low-intramuscular fat (IMF) rabbit lines in plasma metabolites related to liver
Trait Mean SD D
1
HPD
95%2
P
03
r
4
P
r5
Glucose (mg/dl) 141 10.2 −0.90 −6.61, 4.47 0.63 3.38 0.20
Triglycerides (mg/dl) 130 58.6 −43.6 −79.3, −6.86 0.99 19.5 0.91
Cholesterol (mg/dl) 78.4 16.4 −6.78 −16.1, 2.64 0.93 5.47 0.61
Bilirrubin (mg/dl) 0.20 0.11 −0.12 −0.18, −0.06 1.00 0.04 0.99
Total protein (g/dl) 6.81 0.54 0.00 −0.28, 0.31 0.51 0.18 0.12
Albumin (g/dl) 4.36 0.26 0.23 0.07, 0.37 1.00 0.09 0.96
AST (UI/l) 40.6 9.48 1.59 −4.13, 7.23 0.72 3.16 0.29
ALT (UI/l) 69.4 19.6 15.05 3.99, 25.9 1.00 6.52 0.93
ALP (UI/l) 616 111 −99.8 −165, −40.3 1.00 37.1 0.97
AST =aspartate transaminase; ALT =alanine transaminase; ALP =alkaline phosphatase.
1
Median of the marginal posterior distribution of the difference between high and low-intramuscular fat lines.
2
Highest posterior density region at 95% of probability.
3
Probability of the difference being greater than 0 when D >0 or lower than 0 when D <0.
4
Relevant value, proposed as one-third of the standard deviation of the trait.
5
Probability of relevance (probability of the difference being greater than
r
when D >0 or lower than
r
when D <0).
Table 4
Phenotypic correlations of intramuscular fat and perirenal fat
weight with liver weight, lipogenic
1
activities and plasma metabolites
concentrations related to liver in rabbits
Intramuscular fat Perirenal fat weight
Trait
r
p
2
HPD
95%3
P
04
r
p
2
HPD
95%3
P
04
Liver weight 0.28 0.04, 0.51 0.98 0.16 −0.08, 0.42 0.89
G6PDH 0.28 0.02, 0.51 0.97 0.16 −0.11, 0.40 0.88
ME1 −0.05 −0.33, 0.24 0.62 0.34 0.08, 0.57 0.99
FASN 0.38 0.14, 0.60 1.00 0.17 −0.09, 0.43 0.89
Albumin 0.27 0.01, 0.51 0.98 0.35 0.12, 0.57 1.00
Total protein 0.21 −0.06, 0.46 0.94 0.12 −0.14, 0.37 0.82
1
Activities of the lipogenic enzymes glucose-6-phosphate dehydrogenase
(G6PDH), malic enzyme (ME1) and fatty acid synthase (FASN) measured in nmol/
min and g of tissue.
2
Median of marginal posterior distribution of the phenotypic correlation.
3
Highest posterior density region at 95% of probability.
4
Probability of the phenotypic correlation of being greater than 0 when
r
p
>0or
lower than 0 when
r
p
<0.
Martínez-Álvaro, Paucar, Satué, Blasco and Hernández
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Differences in the fat deposition of the high-IMF and low-IMF
lines can be explained by different G6PDH and ME1 lipogenic
activities in liver. Differences between lines were particularly
great (1.51 SD) and relevant for G6PDH, which was the main
lipogenic activity in rabbit liver, in agreement with other
studies in rabbits (Gondret
et al
., 1997 and 2004). We did not
observe differences between lines for FASN activity, although
these results should be taken with caution because of large
HPD
95%
. Both G6PDH and ME1 enzymes generate NADPH
for the support of fatty acid and steroid biosynthesis, G6PDH
by the hexose monophosphate shunt and ME1 by the citric
acid cycle. In a previous study of the lipogenic activities in LD,
semimembranosus proprius
muscle and perirenal fat of
the lines, Martínez-Álvaro
et al
. (2017) observed greater
lipogenic activities in the high-IMF line at 13 weeks, but not
at 9 weeks, in all tissues. Moreover, differences between
lines at 13 weeks were particularly great in the G6PDH
activity of LD. Results after selection for IMF reveal the
important role of G6PDH activity in the genetic variability on
fat deposition in rabbits. Enzyme G6PDH also serves to pro-
duce NADPH which is included in oxidative/antioxidant
metabolism (Ying, 2008). However, the oxidative capacity in
the liver of rabbit lines remains to be analyzed to evaluate its
relationship with G6PDH activity.
Liver lipogenic activities have been previously measured in
breeds with different IMF; however, this is the first work that
studies liver lipogenic activities in animals with the same
genetic origin, divergently selected for IMF. Greater FASN
gene expression in liver has been related to greater IMF in a
comparison between two chicken breeds (Cui
et al
., 2012).
However, breeds can differ in a wide set of traits, which
made difficult to attribute the causes of the differences in
IMF. Several studies show that animals with greater carcass
fat deposition have greater liver weight (Wise
et al
., 1993
and Pond
et al
., 1992 in pigs divergently selected for plasma
total cholesterol) and greater G6PDH, ME1 and FASN acti-
vities in liver (Turkenkopf
et al
., 1980 and Smith and Kaplan,
1980 in fat genotyped Zucker rats). In pigs, Muñoz
et al
.
(2013) observed that selection for decreased backfat thick-
ness at constant IMF was accompanied by a reduction of
FASN expression in liver, suggesting that hepatic lipogenesis
might affect fat partitioning in pigs (Muñoz
et al
., 2013).
Our lines showed normal concentrations of all plasma
metabolites except for ALP, in which both lines showed
concentrations above normal levels for rabbits (Washington
and Van Hoosier, 2012). However, Melillo (2007) suggested
that high plasma concentration of ALP in healthy rabbits is a
common finding, since ALP is the sum of three different
isoenzimes (two isoenzimes produced in the liver and one in
the intestine) with a wide range of variation. Besides,
growing rabbits show particularly high ALP concentrations
caused by its high osteoblastic activity, since ALP is involved
in the precipitation of calcium phosphate in bones (Melillo,
2007). To our knowledge, our results are the first reports
of plasma metabolites in animals selected for IMF.
Circulating plasma concentrations of glucose, triglycerides
and cholesterol are the result of the production and uptake
by lipogenic tissues. We did not find differences between
lines for glucose concentration, although the HPD
95%
of the
difference between lines was large. Low-IMF line had greater
plasma triglycerides and cholesterol concentrations than
high-IMF line in spite of its lower liver lipogenic activity.
A study in rats observed that high plasma concentrations of
triglyceride-rich lipoproteins played a regulation role inhi-
biting hepatic FASN (Lakshmanan
et al
., 1977). In animals
selected for different criterions, it has been observed a
negative relationship between plasma lipids and carcass fat
deposition (Bakke, 1975 selecting for BW gain and carcass
leanness and Pond
et al
., 1992 selecting for plasma choles-
terol, both in pigs). The lower fat deposition of the low-IMF
line suggests that its increased concentration of lipids in
plasma is not taken up by muscles and fat depots in a similar
rate than in the high-IMF line. The release of plasma lipids to
muscle and fat tissues are limited by the activity of the
enzyme lipoprotein lipase, which has been suggested as a
good indicator of lipid deposition in pigs (Allen
et al
., 1976).
Further studies would be necessary to examine the lipo-
protein lipase activity of the IMF lines.
Bilirubin is a subproduct of haemolysis and it is taken up
from plasma by the liver (Wang
et al
., 2006). Low-IMF line
showed relevantly greater plasma concentration of bilirubin
than the high-IMF line. In healthy humans, greater body fat
percentage is related with lower plasma concentration of
bilirubin (Jenko-Praznikar
et al
., 2013). This is explained
because obesity is associated with an increased oxidative
stress and inflammation states, and bilirubin, which has
antioxidant and anti-inflammatory properties, is greatly
consumed in obese individuals (Jenko-Praznikar
et al
., 2013).
Albumin is synthetized in liver and represents the
main part of the total protein concentration in plasma
(Washington and Van Hoosier, 2012). It transports many
plasma metabolites, including bilirubin and free fatty
acids. High-IMF line showed relevantly greater albumin
concentration than low-IMF line, which can indicate greater
transport fluxes of these metabolites in plasma. Although we
did not find difference between lines in total protein, this
result was estimated with a large HPD
95%
.
Plasma concentrations of ALT, AST and ALP enzymes are
used clinically as indicators of liver damage, which was not
the case of none of our lines. High-IMF line showed relevant
greater ALT concentration than the low-IMF line. This
enzyme is involved in the amino acids metabolism (Frayn,
1998). By other side, plasma concentration of ALP was
relevantly greater in the low than in the high-IMF line. We
did not find information about the relationship of IMF with
ALT, AST and ALP plasma concentrations, but pigs with
higher carcass adiposity showed greater ALT, AST and
lower ALP plasma concentrations with respect to leaner
pigs, in a selection experiment for plasma cholesterol (Pond
et al
., 1997).
Intramuscular fat and perirenal fat weight were both
positively correlated with liver weight and lipogenic activities
although the correlations were low. These results suggest
that fat deposition in rabbits, both in muscle and carcass, is
Metabolism of intramuscular fat
5
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partially explained by the liver lipogenic activity. However, all
the correlation estimates showed a wide HPD
95%
and we
cannot make precise statements about their actual values. To
our knowledge, there is no literature about the correlations
between intramuscular and carcass fat and liver lipogenic
activities.
Correlations between IMF and plasma metabolites may
have a particular interest in meat production, because they
could be used as potential biomarkers of IMF. However, we
did not find any strong correlation between IMF and
studied plasma metabolites. Plasma metabolites have been
previously studied as blood indicators of IMF in pigs
(Muñoz
et al
., 2012) and cattle (Adachi
et al
., 1999) with no
significant results. These findings suggest the complex
biological mechanisms involved in the regulation of IMF
deposition, making difficult to find one specific biomarker
strongly correlated to IMF.
Conclusions
Liver plays an important role in the fat deposition of the lines
divergently selected for IMF, high-IMF line showing greater
liver weight and liver lipogenic activities (G6PDH and ME1)
than low-IMF line, particularly for G6PDH. Liver size and liver
lipogenic activities were positively correlated with fat
deposition in muscle (except for ME1) and carcass, although
phenotypic correlations were estimated with low accuracy.
Selection for IMF affected some plasma metabolites related
to liver metabolism, low-IMF line showing greater con-
centration of triglycerides, cholesterol, bilirubin and ALP but
lower concentrations of albumin and ALT than high-IMF line.
Nevertheless, none of these plasma metabolites showed a
strong correlation with IMF.
Acknowledgements
This work was supported by project AGL2014-55921-C2-01-P
from the Spanish National Research Plan. M.M.A. acknowl-
edges a FPI (grant no: BES-2012-052655) from the Economy
Ministry of Spain. The authors thank Federico Pardo for its
technical assistance.
References
Adachi K, Kawano H, Tsuno K, Nomura Y, Yamamoto N, Arikawa A, Tsuji A,
Adachi M, Onimaru T and Ohwada K 1999. Relationship between serum bio-
chemical values and marbling scores in Japanese Black steers. Journal of
Veterinary Medical Science 61, 961–964.
Allen CE, Beitz DC, Cramer DA and Kauffman RG 1976. Biology of fat in meat
animals. North Central Regional Research Publication No 234. University of
Wisconsin, Madison, USA.
Bakke H 1975. Serum levels of cholesterol in lines of pigs selected for rate of gain
and thickness of backfat. Acta Agriculture Scandinavica 25, 14–16.
Ballard FJ, Hanson RW and Kronfield DS 1969. Gluconeogenesis and lipogenesis
in tissue from ruminant and non-ruminant animals. Federation Proceedings 28,
218–231.
Blasco A 2017. Bayesian analysis for animal scientists. Springer, New York, NY, USA.
Blasco A and Ouhayoun J 1996. Harmonization of criteria and terminology in
rabbit meat research. Revised proposal. World Rabbit Science 4, 93–99.
Ciobanu DC, Lonergan SM and Huff-Lonergan EJ 2011. Genetics of meat quality
and carcass traits. In The genetics of the pig (ed. MF Rothschild and A Ruvinsky),
pp. 355–389. CABI Publishing, Oxfordshire, UK.
Cui HX, Zheng MQ, Liu RR, Zhao GP, Chen JL and Wen J 2012. Liver dominant
expression of fatty acid synthase (FAS) gene in two chicken breeds during
intramuscular-fat development. Molecular Biology Reports 39, 3479–3484.
European Commission Directive 2010. Council, E. P. A. E. 2010/63/ EU on the
protection of animals used for scientific purposes. Institute for Health and
Consumer Protection, Ispra, Italy, b7.
Frayn KN 1998. Regulación del metabolismo: una perspectiva humana. Omega,
Barcelona, Spain.
Gondret F, Hocquette JF and Herpin P 2004. Age-related relationships between
muscle fat content and metabolic traits in growing rabbits. Reproduction
Nutrition Development 44, 1–16.
Gondret F, Mourot J and Bonneau M 1997. Developmental changes in lipogenic
enzymes in muscle compared to liver and extramuscular adipose tissues in
the rabbit (oryctolagus cuniculus). Biochemistry and Molecular Biology 117B,
259–265.
Hernández P, Ariño B, Grimal A and Blasco A 2006. Comparison of carcass and
meat characteristics of three rabbit lines selected for litter size or growth rate.
Meat Science 73, 645–650.
Hernández P, Pla M, Oliver MA and Blasco A 2000. Relationships between meta
quality mesurements in rabbits fed with three diets of different fat type and
content. Meat Science 55, 379–384.
Jenko-Praznikar Z, Petelin A, Jurdana M and Ziberna L 2013. Serum bilirubin
levels are lower in overweight asymptomatic middle-aged adults: an early
indicator of metabolic syndrome? Metabolism Clinical and Experimental 62,
976–985.
Kaplan LA, Pesce AJ and Kazmierczak SC 2009. Clinical chemistry: theory,
analysis, correlation, 5th edition. C.V. Mosby, Toronto, Canada.
Lakshmanan MR, Muesing RA, Cook GA and Veech RL 1977. Regulation of
lipogenesis in isolated hepatocytes by triglyceride-rich lipoproteins. Journal of
Biological Chemistry 252, 6581–6584.
Legarra A, Varona L and López de Maturana E 2008. TM: Thershold model.
GenoToul Bioinformatics, Tolouse, France. Retrieved on 7 May 2017 from http://
genoweb.toulouse.inra.fr/~alegarra/tm_folder/
Martínez-Álvaro M, Agha S, Blasco A and Hernández P 2017. Muscle lipid
metabolism in two rabbit lines divergently selected for intramuscular fat. Journal
of Animal Science 95, 2576–2584.
Martínez-Álvaro M, Hernández P and Blasco A 2016. Divergent selection on
intramuscular fat in rabbits: responses to selection and genetic parameters.
Journal of Animal Science 94, 4993–5003.
Melillo A 2007. Rabbit clinical pathology. Journal of Exotic Pet Medicine 16,
135–145.
Muñoz R, Estany J, Tor M and Doran O 2013. Hepatic lipogenic enzyme
expression in pigs affected by selection for decreased backfat thickness at
constant intramuscular fat content. Meat Science 93, 746–751.
Muñoz R, Tor M and Estany J 2012. Relationship between blood lipid indicators
and fat content and composition in Duroc pigs. Livestock Science 148, 95–102.
O’Hea EK and Leveille GA 1969. Lipid biosynthesis and transport in the domestic
chick (
Gallus domesticus
). Comparative Biochemistry and Physiology 30, 149–
159.
Pond WG, Insull W, Mersmann HJ, Wong WW, Harris KB, Cross HR, Smith EO,
Heath JP and Kömüves LG 1992. Effect of dietary fat and cholesterol level on
growing pigs selected for three generations for high or low serum cholesterol at
age 56 days. Journal of Animal Science 70, 2462–2470.
Pond WG, Su DR and Mersmann HJ 1997. Divergent concentrations of plasma
metabolites in swine selected for seven generations for high or low plasma total
cholesterol. Journal of Animal Science 75, 311–316.
Sapp RL, Bertrand JK, Pringle TD and Wilson DE 2002. Effects of selection for
ultrasound intramuscular fat percentage in Angus bulls on carcass traits of
progeny. Journal of Animal Science 80, 2017–2022.
Schwab CR, Baas TJ, Stalder KJ and Nettleton D 2009. Results from six gene-
rations of selection for intramuscular fat in Duroc swine using real-time ultra-
sound. I. Direct and correlated phenotypic responses to selection. Journal of
Animal Science 87, 2774–2780.
Smith PA and Kaplan ML 1980. Development of hepatic and adipose tissue
lipogenesis in the fa/fa rat. International Journal of Biochemistry 11, 217–228.
Martínez-Álvaro, Paucar, Satué, Blasco and Hernández
6
https://www.cambridge.org/core/terms. https://doi.org/10.1017/S1751731117002695
Downloaded from https://www.cambridge.org/core. University of Edinburgh, on 30 Oct 2017 at 09:46:35, subject to the Cambridge Core terms of use, available at
Turkenkopf IJ, Olsen JL, Moray L, Greenwood MRC and Johnson PR 1980.
Hepatic lipogenesis in preobese Zucker rat (40910). Proceedings of the Society
for experimental Biology and Medicine 164, 530–533.
Wang X, Roy Chowdhury J and Roy Chowdhury N 2006. Bilirubin metabolism:
applied physiology. Current Paediatrics 16, 70–74.
Washington IM and Van Hoosier GV 2012. Clinical biochemistry and hemato-
logy. In The laboratory rabbit, guinea pig, hamster, and other rodents
(ed. MA Suckow, KA Stevens and RP Wilson), pp. 57–116. Blackwell Publishing
Professional, Ames, Iowa, USA.
Wise T, Young DL and Pond WG 1993. Reproductive, endocrine and organ
weight differences of swine selected for high or low serum cholesterol. Journal
of Animal Science 71, 2732–2738.
Wood JD, Enser M, Fisher AV, Nute GR, Sheard PR, Richardson RI, Hughes SI and
Whittington FM 2008. Fat deposition, fatty acid composition and meat quality:
a review. Meat Science 78, 343–358.
Ying W 2008. NAD +/NADH and NADP +/NADPH in cellular functions an cell death:
regulation and biological consequences. Antioxidants & Redox Signaling 10, 179–206.
Zhao GP, Chen JL, Zheng MQ, Wen J and Zhang Y 2007. Correlatedresponses to
selection for increased intramuscular fat in a Chinese quality chicken line.
Poultry Science 86, 2309–2314.
Zomeño C, Blasco A and Hernández P 2010. Influence of genetic line on lipid
metabolism traits of rabbit muscle. Journal of Animal Science 88, 3419–3427.
Zomeño C, Hernández P and Blasco A 2011. Use of near infrared spectroscopy
for intramuscular fat selection in rabbits. World Rabbit Science 19, 203–208.
Metabolism of intramuscular fat
7
https://www.cambridge.org/core/terms. https://doi.org/10.1017/S1751731117002695
Downloaded from https://www.cambridge.org/core. University of Edinburgh, on 30 Oct 2017 at 09:46:35, subject to the Cambridge Core terms of use, available at
