Annotation 2

Works Cited

Verbrugghe, Adronie, et al. "The Glucose and Insulin Response to Isoenergetic Reduction of Dietary Energy Sources in a True Carnivore: The Domestic Cat (Felis Catus)." The British journal of nutrition 104.2 (2010): 214-21. ProQuest. Web. 24 Sep. 2014.

The glucose and insulin response to isoenergetic reduction of dietary energy

sources in a true carnivore: the domestic cat (Felis catus)

Adronie Verbrugghe1*, Myriam Hesta1, Stephanie Van Weyenberg1, Georgios A. Papadopoulos1,

Kris Gommeren2, Sylvie Daminet2, Tim Bosmans2, Ingeborgh Polis2, Johan Buyse3 and Geert P. J. Janssens1

1Laboratory of Animal Nutrition, Faculty of Veterinary Medicine, Ghent University, Heidestraat 19, B-9820 Merelbeke, Belgium

2Department of Small Animal Medicine, Faculty of Veterinary Medicine, Ghent University, Salisburylaan 133, B-9820 Merelbeke,

Belgium

3Laboratory of Livestock Physiology, Immunology and Genetics of Domestic Animals, Department of Biosystems, K.U. Leuven,

Kasteelpark Arenberg 30, B-3001 Heverlee, Belgium

(Received 11 August 2009 – Revised 12 January 2010 – Accepted 13 January 2010 – First published online 2 March 2010)

The present study assessed the effect of separate reduction of each energy-delivering nutrient – protein, fat and carbohydrate – on glucose

tolerance and insulin response in a strict carnivore: the domestic cat (Felis catus). Three isoenergetic, home-made diets with the following

energetic distribution, low protein (LP): protein 28% of metabolisable energy; fat 43 %; nitrogen-free extract 29 %; low fat: 47, 27 and 25 %;

low carbohydrate (LC): 45, 48 and 7%, were tested in a 3 £ 3 Latin square design. Nine healthy normal-weight cats were randomly assigned

to each of the diets in a random order at intervals of 3 weeks. At the end of each testing period, intravenous glucose tolerance tests were performed.

Plasma glucose concentrations and area under the glucose curve showed no differences. Area under the insulin curve was lower when cats were fed

the LP diet, and the second insulin peak tended to be delayed when the LC diet was fed. In contrast to other studies, in which energy sources were

elevated instead of being reduced, the present trial contradicts the often suggested negative impact of carbohydrates on insulin sensitivity in

carnivores, and shows that reducing the dietary carbohydrate content below common amounts for commercial foods evokes an insulin-resistant

state, which can be explained by the cats’ strict carnivorous nature. It even points to a negative effect of protein on insulin sensitivity, a finding

that corresponds with the highly gluconeogenic nature of amino acids in strict carnivores.

Carbohydrates: Carnivores: Cats: Energy sources: Insulin resistance

Insulin resistance is a state in which greater than normal insulin

concentrations are required to elicit a quantitatively normal

glucose response in the body, tissues and cells(1). Insulin

resistance and consequent hyperinsulinaemia are associated

with a cluster of abnormalities, such as hypertension, dyslipidaemia

and central obesity, which increase cardiovascular

risk. This cluster is referred to as the metabolic syndrome(1,2).

In individuals who are unable to compensate for reduced insulin

sensitivity, impaired glucose tolerance and overt diabetes

might occur because of the prolonged and increased demand

on b-cells to secrete insulin(1,3).

A critical role for the quantity and quality of dietary carbohydrates

in the pathogenesis of this disorder has been postulated

by the ‘carnivore connection’ theory(1,3,4). During

the Ice Ages, our ancestors consumed high-protein (HP),

low-carbohydrate (LC) diets, and since the brain, fetus and

mammary gland all have specific needs for glucose, metabolic

adaptations were necessary to adapt to low glucose intake.

Therefore, resistance to the glucose-lowering effects of insulin

offered survival and reproductive advantages. The event of the

agricultural revolution augmented the amount of digestible

carbohydrates, and the industrial revolution was responsible

for changing the quality of carbohydrates. These evolutionary

changes in carbohydrates, meaning the introduction of highglycaemic

index foods, can worsen insulin resistance and

can be linked to the development of type 2 diabetes(1,4).

Over the last decades, as in humans, the diet of strictly

carnivorous domestic cats changed from HP, LC prey(5) to

commercial diets, often containing moderate to high amounts

of highly digestible carbohydrates. As in humans, these

dietary changes are held responsible for the recent increase

in incidence of feline insulin resistance and diabetes

mellitus(3,6).

Several human studies(7 – 10) as well as rodent studies

(rats(11) and mice(12)) demonstrated that glucose tolerance

and insulin sensitivity could benefit from HP, LC diets. In

healthy cats, high-carbohydrate (HC) diets are suggested to

impair glucose tolerance(13). However, the hypothesis that

HC diets lead to b-cell exhaustion was contradicted by

Slingerland et al. (14), since feeding a HC diet resulted in

* Corresponding author: Adronie Verbrugghe, fax þ32 92647848, email [email protected]

Abbreviations: AUC, area under the curve; HC, high carbohydrate; HF, high fat; HP, high protein; IVGTT, intravenous glucose tolerance tests; LC, low

carbohydrate; LF, low fat; LP, low protein.

British Journal of Nutrition (2010), 104, 214–221 doi:10.1017/S0007114510000358

q The Authors 2010

British Journal of Nutrition

increased glucose-induced insulin secretion during hyperglycaemic

glucose clamps. Yet, this could be the first step

towards b-cell exhaustion, but long-term consequences were

not investigated. It can, however, not be disclaimed that HC

diets might be an indirect risk factor for diabetes by promoting

obesity. Hoenig et al. (15) suggested that cats with the same

energetic intake are more prone towards obesity and insulin

resistance when fed a low-protein (LP), HC diet in comparison

with a HP, LC diet. In contrast, high-fat (HF), LC diets were

also shown to impair glucose tolerance(16,17) as well as to

induce weight gain(16).

A plausible explanation for the lack of agreement among

these studies is that the effect of increasing one energy

source (1) was often confounded with the decrease of other

energy sources; or (2) often meant an increase on top of an

already high level of this energy source.

The present study deals with these aspects by applying a

pairwise reduction of one energy source, and therefore enables

the identification of the separate effect of each energy source

by looking at the effect of its reduction to minimal amounts.

This method has been shown to be effective in demonstrating

single energy source effects on metabolism in poultry(18).

Material and methods

Animals and housing

Six mixed-breed and three European shorthair cats, five

intact females and one intact and three neutered males, were

employed in the present study. All cats were aged between

3 and 10 years, and had a mean body weight of 3·67 kg

(range 2·50–5·24 kg). Body condition score was determined

using a five-point body condition scoring system(19). Nonobese

cats with a body condition score of 2·5/5 to 3·5/5

were used. All cats were healthy and were not given any

medication at the time of the study; none had prior medical

problems. Cats were divided into three groups based on sex

and body weight and were housed individually in separated

indoor cages during the trial. For 2 h a day, the cats were

allowed to play in their usual group cages. At that time,

the cats had no access to the food, but water was available

ad libitum.

Diets and feeding

Three isoenergetic home-made diets were tested: a LP, a lowfat

(LF) and a LC diet. To produce the test diets, cooked and

ground chicken breast was mixed with liquid chicken lard and

maize starch (CstarGel 03 401, native maize starch; Cargill,

Sas van Gent, The Netherlands). The same ingredients were

used in different quantities in order to create pairwise changes

in macronutrient content (Table 1). The LP diet differed from

LF and LC diets only by isoenergetic substitution of protein

for fat and protein for carbohydrate, respectively. The LF

diet differed from the LC diet by isoenergetic substitution of

fat for carbohydrate. The three test diets contained no dietary

fibre, and had similar mineral concentrations and physical

structure. A tailor-made vitamin and mineral premix (Institute

for Physiology, Physiological Chemistry and Nutrition,

Ludwig-Maximilians-University Mu¨nchen, Oberschleibheim,

Germany) was added to balance the diet. The three diets

were analysed for proximate components (Table 1), and

energetic distributions were as follows: LP diet: protein

28 %, fat 43% and nitrogen-free extract 29 %; LF diet:

protein 47 %, fat 27% and nitrogen-free extract 25 %; LC

diet: protein 45 %, fat 48% and nitrogen-free extract 7%.

Cats were offered only one meal daily. The amount of food

corresponded with the cats’ individual maintenance energy

requirement (375 kJ/kg0·75)(20), and was adjusted to maintain

body weight. The food was available all day, except for the

2 h playtime. All cats had free access to water at all times.

Fresh water was supplied daily.

Experimental design

Before being entered into the study, each cat was physically

examined; body weight and body condition score were

recorded and a blood sample was drawn from the jugular

Table 1. Composition of the test diets

LP LF LC

Ingredients (%)

Chicken fillet 66·3 81·0 86·1

Chicken lard 11·4 4·3 8·3

Maize starch* 20·7 14·1 5·0

Vitamin/mineral premix† 1·5 0·6 0·6

Nutrients on DM (%; by Weende analysis)

Crude protein 36·1 54·5 60·2

Diethyl ether extract 23·2 13·1 26·6

Crude fibre 0·4 0·4 0·6

Crude ash 3·1 4·3 4·1

NFE‡ 37·2 27·7 8·5

Starch 26·0 27·1 3·3

TDF 1·9 1·8 3·7

ME (kJ/100 g DM)§ 1938 1762 2025

Nutrients on energy basis (g/MJ ME)

Crude protein 18·7 32·0 30·3

Diethyl ether extract 12·0 7·7 13·4

Crude fibre 0·2 0·2 0·3

Crude ash 1·6 2·5 2·1

NFE 19·2 16·2 4·3

Starch 13·4 15·9 1·7

Amino acid content on DM (%)

Asp 2·8 3·8 3·9

Thr 1·4 2·2 2·3

Ser 1·1 1·7 1·9

Glu 4·4 6·3 6·5

Gly 1·4 2·2 2·2

Ala 2·0 3·0 3·1

Val 1·7 2·5 2·6

Ile 1·8 2·6 2·6

Leu 2·7 4·0 4·1

Tyr 1·5 2·2 2·4

Phe 1·4 2·0 2·1

His 1·1 1·7 1·8

Lys 3·0 4·4 4·7

Arg 2·0 3·0 3·0

LP, low protein; LF, low fat; LC, low carbohydrate; NFE, nitrogen-free extract; TDF,

total dietary fibre; ME, metabolisable energy.

* Native maize starch (CstarGel 03 401, Cargill).

†Provides: LP diet: Ca 14·10 %, P 3·41%, K 8·4%, Mg 0·3 %, Na 0·34 %, Fe 0·31 %,

Zn 0·29 %, Mn 0·038 %, Cu 0·014 %, iodine 0·006 %, vitamin A 45mg/g, vitamin D3

0·31mg/g, vitamin E 3·25 mg/g, vitamin B1 0·3 mg/g, vitamin B2 0·2 mg/g, taurine

75 mg/g; LF and LC diets: Ca 27·70%, Fe 0·44%, Zn 0·56%, Mn 0·057 %, iodine

0·013 %, vitamin A 67·5mg/g, vitamin D3 0·56mg/g, vitamin E 5·65 mg/g, vitamin B1

0·53 mg/g, vitamin B2 0·28 mg/g, taurine 140 mg/g.

‡ Derived by subtracting crude protein, diethyl ether extract, crude fibre and crude

ash from the DM content.

§ Estimated by using a four-step calculation(27).

Isoenergetic reduction of energy sources 215

British Journal of Nutrition

vein after a 12 h fast for complete blood count and serum biochemistry.

For 4 weeks preceding the trial, all cats were fed a

standard commercial maintenance diet (Bento Kronen Torka

chicken-turkey; Versele-Laga, Deinze, Belgium), before

being randomised to one of the three groups. Each group of

cats was assigned to each of the three test diets in a random

order at intervals of 3 weeks. This way, the test diets were

examined in a 3 £ 3 Latin square design.

Absolute food intake was measured each day throughout the

study, and daily energy consumption was calculated. Body

weight was recorded weekly.

To determine the effect on glucose and insulin metabolism,

intravenous glucose tolerance tests (IVGTT) were performed

at the end of each testing period. Hence, at least 20 h before

the IVGTT, cats were anaesthetised with buprenorphine (Temgesic,

Schering-Plough n.v., Heist-Op-Den-Berg, Belgium),

10mg/kg intravenous, followed by propofol (Propovet,

Abbott Lab, Leuven, Belgium), 6–7 mg/kg to effect, intravenous,

and a 20G, 8 cm intra venous catheter (Leaderflex,

Vygon, E ´ couen, France) was placed in a jugular vein for

glucose administration and blood sampling. The catheter was

flushed twice daily with 1ml heparinised saline (50 IU

heparin/ml in 0·9% NaCl solution) to maintain patency.

Amoxicillin (Clamoxyl LA, GlaxoSmithKline n.v., Genval,

Belgium), 15 mg/kg subcutaneous, was administered once at

the time of catheter placement. The IVGTT was performed

between 09.00 and 13.00 hours after a 12 h fast. Glucose,

0·5 g/kg (Glucose Sterop 500 mg/ml, Laboratoria Sterop

n.v., Brussels, Belgium), was administered via the jugular

vein catheter over 30–45 s, followed immediately by 1ml

of saline solution to flush the catheter(21). Blood samples

were collected from the jugular catheter(22) before (0 min)

and 2, 5, 10, 15, 30, 45, 60, 90 and 120 min after glucose

administration(21).

In addition, effects on lipid as well as protein metabolism

were evaluated by the analysis of total cholesterol, TAG,

NEFA, urea and creatinine, respectively. The effect of

energy-delivering nutrients on plasma leptin concentrations

was also investigated, since leptin also influences carbohydrate

and fat metabolism(23).

At time zero, blood samples were collected in tubes

containing lithium heparin for the determination of plasma

leptin and NEFA concentrations, and serum tubes were used

to determine basal serum total cholesterol, TAG, urea and

creatinine concentrations. At each time interval, blood

samples were collected in tubes containing NaF for the

determination of plasma glucose and in serum tubes for the

determination of serum insulin concentrations. Plasma and

serum were removed by centrifugation and stored at 2208C

until assayed. The experimental protocol of the present

study was approved by the Ethical Committee of the Faculty

of Veterinary Medicine, Ghent University, Belgium (EC

2006/029), and was in accordance with the institutional and

national guidelines for the care and use of animals.

Analytical methods