Postprandial changes in metabolic pathways induced by increasing protein at the expense of carbohydrates.

<p>Higher protein content in the diet increased protein oxidation and urea production. First regulated step of gluconeogenesis was up-regulated (PEPCK), but not the last one (G6PC1). With lower CHO content in the diet, glucose oxidation and liver glycogen content decreased, concomitantly with glycolytic genes expression (GK, LPK), inducing lower lipogenesis (ACC, FAS). Stable fat content in the diet caused no changes in β-oxidation (CPT1, ACOX1, βHAD), that was only transiently increased only after the first day of a HP diet.</p

Body weight and postprandial plasma hormones level (insulin, glucagon and insulin/glucagon (I/G) ratio), glucose level (in arterial, portal and vena cava blood) and liver glycogen measured 2 h after the beginning of 4 g of NP or HP meal after 1, 3, 6 or 14 days of adaptation o NP or HP diet (NP1, NP3, NP6, NP14 and HP1, HP3, HP6, HP14; respectively).

<p>Data are expressed as means±SEM (n = 5).</p><p>*Significantly different between groups at the same day of feeding P<0.05.</p

Composition of NP and HP diets.

<p>*¹IDI;Arras, France;</p>2<p>Cerestar, Haubourdin, France;</p>3<p>Eurosucre, Paris, France;</p>4<p>Bailly SA, Aulnay-sous-bois, France;</p>5<p>ICN Biochemicals, Ohio, USA (see Reeves et al. 1993 for composition);</p>6<p>Medias filtrants Durieux, Torcy, France. P/E, G/E, L/E: percentage of diet energy provided by protein, carbohydrates and lipids, respectively.</p

Postprandial macronutrient balance.

<p>Macronutrient balance was assessed during the 4 h period for total metabolism after the intake of a calibrated meal consisting of 4 g of an adequate diet. The balance was calculated as the difference between absorbed macronutrient and the macronutrient oxidised. The rats were fed normal protein (NP) diet for a week and then switched to a HP diet for 0, 1, 3, 6 and 14 days (NP, HP1, HP3, HP6, HP14, respectively). Data are means±SEM. Significance was determined by one-way ANOVA (P<0.05).</p

Primer sequences used for real- time rt-PCR.

<p>The forward and reverse primers were designed using primer express software (Applied Biosystems). The following abbreviations were used: ACC, acetyl-CoA carboxylase, ACOX1, acyl-CoA oxidase, CPT1, carnityne acyltransferase 1, liver isoform, CPT1b, carnityne acyltransferase 1b, muscle isoform, FAS, fatty acid synthase, G6PC1, glucose-6-phosphatase catalytic subunit, GK, glucokinase, GS1, glycogen synthase, muscle isform, GS2, glycogen synthase, liver isoform, GYG, glycogenin, βHAD, β-hydroxyacyl-Coenzyme A dehydrogenase, HSL, hormone sensitive lipase, L-PK, liver pyruvate kinase, PEPCK, phosphenolpyruvate carboxykinase.</p

Postprandial hepatic gene expression profiles for carbohydrate metabolism.

<p>Gene expression was determined for the key enzymes involved in: (a) glycolysis (L-PK, GK), (b) gluconeogenesis (PEPCK, G6PC1) and (c) glycogenogenesis (glycogenin, GS) 2 h after intake of a calibrated meal consisting of 4 g of NP or HP diet for rats fed previously a normal or a high protein diet for 1, 3, 6 and 14 days (NP1, NP3, NP6, NP14 and HP1, HP3, HP6, HP14, respectively). mRNA levels were measured by real time RT-PCR and expressed in comparison to the reference gene (18S). Data are expressed as means±SEM relatively to NP1 (n = 5). The statistical differences (P<0.05) are shown at the bottom of each graph (two-way ANOVA test).</p

Postprandial gene expression profiles of enzymes encoding glycogen synthesis and fatty acids oxidation pathways in muscles (gastrocnemius and soleus), gluconeogenesis in kidney and fatty acids synthesis and oxidation in adipose tissues (epidydymal and retroperitoneal) for rats fed normal protein diet for two weeks (NP 14) vs. high protein diet for two weeks (HP14), following one week of acclimatisation on NP diet.

<p>Measurements were made 2 h after intake of a 4 g calibrated meal of NP or HP diet. Data are means*10⁵±SE expressed as relative values normalised with 18S.</p><p>*P<0.05.</p

Nutrient oxidation.

<p>Fasting (a) and postprandial (b) nutrient oxidation measured by indirect calorimetry during 4 h period in the fasted state or after the intake of a calibrated meal consisting of 4 g of NP or HP diet for rats previously fed normal protein diet for a week and then switched to a high protein diet for 0, 1, 3, 6 and 14 days (NP, HP1, HP3, HP6, HP14 respectively) Data are means±SEM (one-way ANOVA). Different letters on the graph indicate significant difference (P<0.05).</p

Impairment of extracellular sugar detection in GLUT2-expressing tissues in transgenic mice.

<p>Effect of a glucose-rich diet on gene expression in liver (A) and adipose tissue (C). Transgenic (Tg) and wild-type (WT) mice were fasted for 48 h and refed for 15 h before liver and epididymal fat pad biopsies. Levels of mRNA were analyzed by real time PCR. Values are presented as means±S.E.M. (n = 3 to 5 mice/group). Statistical differences between refed and fasted mice are indicated by *P<0.05, **P<0.01, and ns non significant. B: GLUT2 protein levels in total membrane preparations from the liver of mice fed a glucose-rich diet for five days. D: Blood glucose concentrations during an insulin tolerance test in wild-type and transgenic mice. Values are presented as means±S.E.M (n = 8 to 10 mice/group).</p

Urinary excretion in mice after impairment of extracellular sugar detection. Metabolic and electrolyte levels in 24-h urine samples from wild-type (WT) or transgenic mice fed a standard or a glucose-rich diet. Values are presented as means±S.E.M. (n = 6 per group). Statistical differences between wild-type and transgenic mice are indicated as *P<0.05 ns non significant, nd indicates not determined.

<p>Urinary excretion in mice after impairment of extracellular sugar detection. Metabolic and electrolyte levels in 24-h urine samples from wild-type (WT) or transgenic mice fed a standard or a glucose-rich diet. Values are presented as means±S.E.M. (n = 6 per group). Statistical differences between wild-type and transgenic mice are indicated as *P<0.05 ns non significant, nd indicates not determined.</p