Figure 1.
Expression of mT2R138, mT2R108 and α-Gustducin (Gust) mRNA in the mouse Gastrointestinal Tract.
mRNA levels were analyzed in the different regions of the GI tract with qRT-PCR and normalized to β-actin levels in each tissue (A–C). Relative quantities were determined using the comparative ΔΔCt method. Each cDNA sample was amplified in duplicate and all data are expressed as the mean ± S.E.M. T2R138 expression was very low in the stomach compared to the small and large intestine (A), whereas T2R108 is more abundant in the stomach compared to the intestine (B). α-Gustducin is more abundant in the stomach and colon compared to the other regions (C). D: Single bands of the predicted size were found for each primer, in all GI segments analyzed as well as in STC1 cells or the tongue, which served as positive controls. No signal was detected with any of the primer in 3T3 cells, which were used as negative control. A:antrum, C:corpus, D:duodenum, J: jejunum, I: ileum, PC: proximal colon, DC: distal colon, STC1: STC 1 cells, BA: β actin; CTR, control; * the tongue is shown as control for α-gustducin.
Figure 2.
Distribution of T2R138 Immunoreactivity in the Gastrointestinal Tract.
Confocal images. T2R138 immunostaining in cells (arrows) of the tongue (A) and in isolated cells (arrows) along the GI tract (B, D, E). B: T2R138 immunoreactive cell in the jejunum. C: Lack of specific staining in a section incubated with T2R138 antibody pre-adsorbed with the antigen against which the antibody was raised. D and E: T2R138 immunoreactive cells in the proximal (D) and distal (E) colon. F: high magnification of a T2R138 immunoreactive cell of the colon showing the granular staining concentrated toward the base of the cell. Calibration bar: 20 µm in A–E, 10 µm in F.
Figure 3.
Cells Expressing T2R138 in the Gut.
Confocal images of mouse ileum showing colocalization of T2R138 (green) (A) with α-Gustducin (red) (B) immunoreactivity; C: shows overlay of both T2R138 and α-Gustducin immunoreactivity. D–I: Confocal images showing colocalization of T2R138 (green) (D, G) with chromogranin A, a marker of enteroendocrine cells (red) (E, H) and overlay of both immunoreactivities in the same cells (F,I) in the ileum (D–F) and distal colon (G–I). Calibration bar: 20 µm.
Figure 4.
TR2138, T2R108 and α-Gustducin (Gust) Regulation by Fasting/Re-feeding in the Stomach.
mRNA levels for each transcript were analyzed by qRT-PCR and normalized to β-actin. T2R138, T2R108 and α-gustducin mRNA levels were markedly decreased by fasting (82%, 53% and 37%, respectively compared to controls) and restored by re-feeding. *p<0.05, **p<0.01 vs. fasted. CTR, control; F, fasted; F/R, re-feeding following fasting.
Figure 5.
Effect of a Low Cholesterol (LE) Diet on T2R138, T2R108 and α-Gustducin (Gust) Expression.
qRT-PCR analysis shows that T2R138 mRNA is significantly up-regulated in the duodenum (DUO) and jejunum (JEJ), but not the ileum (IL), proximal colon (PC) and distal colon (DC), following 7 days of a cholesterol lowering diet. α-gustducin mRNA is also significantly increased in the duodenum. *p<0.05, **p<0.01 vs. control. By contrast, T2R108 mRNA levels were not affected by this diet in any regions of the gut.
Figure 6.
Effect of a High Fat Diet on T2R138, T2R108 and α-Gustducin (Gust) Expression in the Ileum and Colon.
qRT-PCR analysis shows that T2R138 mRNA and α-gustducin mRNA levels are significantly (*p<0.05; **p<0.01) up-regulated in the colon, but not ileum by a long term (8 weeks) high fat (45% and 60%) diet compared to low (10%) fat diet. By contrast, T2R108 mRNA was not affected by this diet.