Scientific Poster presented by WALTHAM's David Allaway at the Experimental Biology Conference 2017 in Chicago, USA

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D Allaway, G Thomas, R Staunton & P Watson, WALTHAM Centre for Pet Nutrition, Melton Mowbray, Leics. UK LE14 4RT
Study Objectives
• To identify changes in plasma metabolite
pools following a reduction in dietary Met,
whilst maintaining total sulphur amino acids
with cysteine (Cys)
• To determine whether additional choline
would support Met functions at the lower
dietary Met:Cys (Cys in diet as cystine)
.
Summary
• The study highlights wide-ranging short-term metabolic effects
of, and adaptations to, a change in dietary Met:Cys and choline
• The Met pool reduced with a lower dietary Met:Cys but was
supported by choline and adaptations after 2 weeks of diet
feeding
• Longer term consequences of the metabolic changes require
elucidation before recommending a restricted Met diet for dogs
Metabolomics Reveals Rapid Alterations and Adaptation in Canine Plasma in
Response to Changes in Dietary Methionine:Cystine and Choline
Material & Methods
Study design: A 3-way crossover design with
18 dogs (Labrador retriever) randomised into 6
rotation groups. Each dog was fed each of the
three diets for a period of 2 weeks, with a 6-7
day washout between diets.
Each dog was fed 1 meal daily (100% MER)
Blood samples taken 20 mins pre- and 1hr and 2 hr post diet on each sampling day
All housing and study procedures complied with the Animals (Scientific Procedures) Act 1986
Figure 1 Study design schematic
Diets: Semi-purified experimental diets
formulated to be isonitrogenous / isocaloric and
differing only in S amino acid ratio and choline
content. Control diet A was fed in a 3 week
baseline phase and in the washout phases.
Metabolic profiling: Metabolic profiling used
UHPLC/MS/MS and GC/MS (Metabolon Inc.).
Statistical analysis: Univariate linear mixed
effect models were fitted for 472 metabolites
from 321 samples. Fixed effects were
interactions of diet, time relative to meal, and
days on diet, with covariates of age and gender,
and random effects were diet nested in dog.
Pairwise comparisons between fixed effects
levels were significant at adjusted p-value <0.05
(Benjamini & Hochberg, 1995).
Figure 3 An illustration of the data-
driven observations made within the
context of methionine cycle metabolism
Results & Interpretation
Diet Met Cys Choline
A: Control 100% 100% 100%
B: Low Met:Cys 40% 163% 100%
C: Low Met:Cys & 2x Choline 40% 163% 200%
Figure 2 Fasted plasma metabolite fold-change
relative to control diet (Diet A), on day 2 and
day 14 after a diet change (left and right
symbols respectively). Metabolites are grouped
by metabolic pathway ontology. Metabolites
have at least one significant fold-change from
control (red).
1. Betaine increased rapidly with low Met:Cys, and
rose further with 2 weeks feeding or with
additional choline
2. After 2 weeks, elevated GAA and creatine
indicate that the dominant methyl-demanding
pathway may be augmented by increased flux
through the methionine cycle
3. Whilst dietary choline may support Met in the
acute phase, adaptation within two weeks re-
establishes fasted Met when fed a low Met:Cys
diet
4. FIGLU increased with low Met:Cys, and further
with additional choline, potentially as a
consequence of alterations in 1C partitioning
5. Increased deoxycarnitine (γ-butyrobetaine) and
carnitine pools at all time points, enhanced by
choline on day 2 suggest a change in carnitine
synthesis and/or transport
6. The acute increase in short-chain acylcarnitines
suggests a disturbance in BCAA metabolism in
response to choline
7. Additional choline resulted in increased TMAO
production
8. Specific lipids decreased after 2 weeks
adaptation to low Met:Cys, indicating reduced
PEMT activity, with additional choline having
variable effects
Introduction
• Dietary methionine (Met) restriction may provide health and longevity
benefits
• In susceptible dogs, dietary Met too low for sufficient taurine production
has been implicated in the development of dilated cardiomyopathy
• It was hypothesised that some health benefits of restricted Met diets in
dogs may be possible if taurine production was maintained through
provision of cysteine (Cys) or choline, which can support the Met cycle
Metabolic Pathway Ontology
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4
5 6 7
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