Etude des conséquences à long terme de l'exposition in utero à un diabète maternel sur la fonction rénale et la pression artérielle chez le rat (implication du rein et du système vasculaire)

PARIS7-Bibliothèque centrale (751132105) / SudocSudocFranceF

Body weight gain and food intake for <i>MCT1</i>^+/+ and <i>MCT1</i>^+/− mice fed high fat diet.

<p>(<b>A</b>) Evolution with time of body weight (<b>B</b>) Cumulative body weight gain (<b>C</b>) Weekly measured food intake (<b>D</b>) Cumulative food intake. Results represent mean ± SEM, n = 10–11. Filled circles, <i>MCT1</i>^+/+ mice; open circles, <i>MCT1</i>^+/− mice. The asterisk represents a statistically significant difference between <i>MCT1</i>^+/+ and <i>MCT1</i>^+/− mice evaluated using a Student’s <i>t</i>-test for each pair of data under the dotted line. *<i>P</i><0.05.</p

Fat mass accumulation and distribution in <i>MCT1</i>^+/+ and <i>MCT1</i>^+/− mice fed a normal chow or high fat diet.

<p>(<b>A</b>) Fat mass changes in <i>MCT1</i>^+/+ and <i>MCT1</i>^+/− mice fed a normal chow or a high fat diet for different periods of time. Results are mean ± SEM, n = 4-9. Asterisks represent a statistically significant difference between <i>MCT1</i>^+/+ and <i>MCT1</i>^+/− mice evaluated using a Student’s <i>t</i>-test. *<i>P</i><0.05, ***<i>P</i><0.001. +/+, <i>MCT1</i>^+/+; +/-, <i>MCT1</i>^+/−; NC, normal chow; HFD, high fat diet; 2 m, 2 months; 9 m, 9 months. (<b>B</b>) Comparative histological analysis of liver and distinct adipose tissues of <i>MCT1</i>^+/+ and <i>MCT1</i>^+/− mice fed a high fat diet for 11 months. ScWAT, subcutaneous white adipose tissue; PgWAT, perigonadal white adipose tissue; IgWAT, inguinal white adipose tissue. Calibration bar, 100 µm.</p

Glucose homeostasis in <i>MCT1</i>^+/+ and <i>MCT1</i>^+/− mice fed a normal chow or high fat diet.

<p>(<b>A</b>) Intraperitoneal glucose tolerance test in <i>MCT1</i>^+/+ and <i>MCT1</i>^+/− mice fed with a normal chow (left panel) or with a high fat diet (right panel) for 8 weeks. Inset: Quantitative analysis of integrated glycemic responses (as area under the curve or AUC). Filled bar, <i>MCT1</i>^+/+ mice. Open bar, <i>MCT1</i>^+/− male mice (<b>B</b>) Intraperitoneal insulin tolerance test in <i>MCT1</i>^+/+ and <i>MCT1</i>^+/− mice fed with a normal chow (left panel) or with a high fat diet (right panel) for 8 weeks. Filled circles, <i>MCT1</i>^+/+; open circles, <i>MCT1</i>^+/−. Results are mean ± SEM, n = 5–9. Asterisks represent a statistically significant difference between <i>MCT1</i>^+/+ and <i>MCT1</i>^+/− mice evaluated using a Student’s <i>t</i>-test. *<i>P</i><0.05, **<i>P</i><0.01, ***<i>P</i><0.001.</p

Gene targeting strategy to produce the <i>MCT1</i> knockout/ß-galactosidase knockin mouse.

<p>(<b>A</b>) Structure of the <i>MCT1</i> wildtype allele, the targeting vector, and the resulting <i>MCT1</i> knockout/ß-galactosidase knockin allele showing the targeted locus. Exons are represented as grey boxes except exon 1 that contains the translation initiation codon (ATG). The <i>mct1</i> gene sequence (640 bp) replaced by the fused LacZ/Neo gene sequence is indicated in red. TK, thymidine kinase gene for negative selection with Gancyclovir ; ß-Gal, the LacZ gene sequence coding for the enzyme ß-galactosidase ; Neo, neomycin resistance gene for positive selection with G418. (<b>B</b>) Southern blot performed with a 900 bp probe (green in A) on selected embryonic cell (ES) DNA digested with NdeI (Nd). ES 1 and 2 cell populations are showing a 7200 nt band corresponding to the wildtype (WT) allele and a 8800 nt band corresponding to the modified knockin (KI) allele due to a double recombination event. ES 3 and 4 cell populations did not undergo the double recombination event and only exhibit the 7200 nt band. (<b>C</b>) PCR genotyping strategy. PCR made with the primers MCT1 Fo, MCT1 Re and ßGal Re (see localization in A) present in the same reaction produces a fragment of 538 nt for wildtype (WT) animals, but also an additional fragment of 422 nt for heterozygote (HE) animals. (<b>D</b>) Histochemical detection of ß-galactosidase activity in selected tissues of <i>MCT1</i>^+/LacZ mice. The blue color in cells reflects the presence of the ß-galactosidase enzyme and therefore the <i>mct1</i> promoter activity (arrows). Tissue counterstaining was done with nuclear fast red dye. EDL, <i>extensor digitorum longus</i>.</p

Locomotor activity and indirect calorimetry for <i>MCT1</i>^+/+ and <i>MCT1</i>^+/− mice fed a normal chow or high fat diet.

<p>(<b>A</b>) Vertical and horizontal movements of <i>MCT1</i>^+/+ and <i>MCT1</i>^+/− mice under normal chow. (<b>B</b>) Vertical and horizontal movements of <i>MCT1</i>^+/+ and <i>MCT1</i>^+/− mice under high fat diet. Filled bars, <i>MCT1</i>^+/+ mice; open bars, <i>MCT1</i>^+/− mice. (<b>C</b>) O₂ consumption and CO₂ production of <i>MCT1</i>^+/+ and <i>MCT1</i>^+/− mice fed a normal chow. (<b>D</b>) O₂ consumption and CO₂ production of <i>MCT1</i>^+/+ and <i>MCT1</i>^+/− mice fed a high fat diet. V<i>O₂</i>, oxygen consumption ; V<i>CO₂</i>, carbon dioxide production. Filled bars, <i>MCT1</i>^+/+ mice; open bars, <i>MCT1</i>^+/− mice. Results represent mean ± SEM, n = 5-8. Asterisks represent a statistically significant difference between <i>MCT1</i>^+/+ and <i>MCT1</i>^+/− mice evaluated using a Student’s <i>t</i>-test. **<i>P</i><0.01.</p

MCT1 mRNA and protein expression levels in tissues from <i>MCT1</i>^+/+ vs. <i>MCT1</i>^+/− mice and comparative histological analysis.

<p>(<b>A</b>) Quantitative RT-PCR analysis of relative MCT1 mRNA levels in tissues collected from <i>MCT1^+/</i>⁻ and <i>MCT1^+/+</i> mice. Values are expressed as percentage and represent the mean normalized MCT1 mRNA level in <i>MCT1^+/</i>⁻ mouse tissue compared to the mean normalized MCT1 mRNA level in <i>MCT1^+/+</i> mouse tissue ± SEM, n = 6. ß-actin mRNA was used as reference. Statistical analysis was performed using the Student’s t-test. Asterisks indicate a significant difference between heterozygote and wildtype tissues. *<i>P</i><0.05, ** <i>P</i><0.01. (<b>B</b>) Representative western blot performed on protein extracts from selected tissues of <i>MCT1^+/+</i> and <i>MCT1^+/</i>⁻ mice. ß-actin was used as reference for quantification. (<b>C</b>) Quantitative analysis of MCT1 protein expression. Values are expressed as percentage, and represent the mean normalized MCT1 protein level of <i>MCT1^+/</i>⁻ mice compared to the mean normalized MCT1 protein level for <i>MCT1^+/+</i> mice ± SEM, n = 6. Statistical analysis was performed using the Student’s <i>t</i>-test. Asterisks indicate a significant difference between heterozygote and wildtype tissues. *<i>P</i><0.05, ** <i>P</i><0.01. (<b>D</b>) Comparative histological analysis of selected tissues from <i>MCT1^+/+</i> and <i>MCT1^+/</i>⁻ mice. Sections from brain (cortex), heart, skeletal muscle (mixed type), liver, and white adipose tissue (WAT) of both <i>MCT1^+/+</i> and <i>MCT1^+/</i>⁻ mice were stained with hematoxylin/eosin and examined under light microscopy at 40x magnification. Calibration bar, 100 µm.</p

Plasma biochemical profile of MCT1^+/+ and MCT1^+/− mice fed a normal chow or a high fat diet.

<p>Results are expressed as mean ± SEM; values within parentheses indicate number of mice. *<i>P</i><0.05, **<i>P</i><0.01, ***<i>P</i><0.001 vs. <i>MCT1^+/+</i> on the same diet with two-way ANOVA followed by Bonferroni post hoc test. HDL, High density lipoprotein; LDL, Low density lipoprotein; FFA, Free fatty acids; LDH, Lactate dehydrogenase; ALAT, Alanine aminotransferase; ASAT, Aspartate aminotransferase; HOMA, Homeostasis model assessment.</p

Body, liver and white adipose tissues mass of HFD fed mice.

<p>Results are expressed as mean ± SEM ; values within parentheses indicate number of mice. *<i>P</i><0.05, **<i>P</i><0.01, ***<i>P</i><0.001 by Student’s <i>t</i>-test. ScWAT, Subcutaneous white adipose tissue; PgWAT, Perigonadal white adipose tissue; IgWAT, Inguinal white adipose tissue.</p