Coffea arabica bean extract inhibits glucose transport and disaccharidase activity in Caco-2 cells

Abstract

The major constituents of Coffea arabica (coffee), including caffeine, chlorogenic acid and caffeic acid, exhibit antihyperglycemic properties in in vitro and in vivo models. However, whether Coffea arabica bean extract (CBE) regulates glucose uptake activity and the underlying mechanisms involved remain unclear. The aim of the present study was to examine the effects of CBE on glucose absorption and identify the mechanisms involved using an in vitro model. The uptake of a fluorescent glucose analog into Caco-2 colorectal adenocarcinoma cells was determined. The expression levels of sodium glucose co-transporter 1 (SGLT1) and glucose transporter 2 (GLUT2) were evaluated. In addition, glycoside hydrolase enzyme activity was investigated. It was observed that CBE inhibited disaccharidase enzyme activity. Furthermore, CBE exerted an inhibitory effect on intestinal glucose absorption by downregulating SGLT1- and GLUT2-mediated 5' AMP-activated protein kinase phosphorylation and suppressing hepatocyte nuclear factor 1α expression. These data suggest that CBE may attenuate glucose absorption and may have potentially beneficial antihyperglycemic effects in the body; however, the mechanisms underlying the effects of CBE must be elucidated through further investigation.

Keywords: 5' AMP-activated protein kinase; CBE; GLUT2; SGLT1; glucose uptake; hepatocyte nuclear factor 1α.

Figure 1

Representative high-performance liquid chromatography chromatogram of Coffea arabica bean extract at an absorption wavelength of 280 nm. mAU, milli-arbitrary units.

Figure 2

Effect of CBE and its constituents on glucose transport. (A) Effect of treatment with 0.1-1,000 µg/ml CBE and its constituents at a molar ratio concentration, including chlorogenic acid (29.62 µg/ml), caffeine (5.11 µg/ml), caffeic acid (3.16 µg/ml), ferulic acid (1.54 µg/ml), as well as the SGLT1 inhibitor phlorizin (0.5 mM) and GLUT2 inhibitor phloretin (1 mM) for 4 h at 37˚C on glucose transport. (B) Viability of Caco-2 cells after exposure to CBE, chlorogenic acid, caffeine, caffeic acid, ferulic acid, phlorizin and phloretin. Values are presented as the mean ± the standard error of the mean (n=6). ^*P<0.05, ^**P<0.01 vs. control. CBE, Coffea arabica bean extract; RFUY, relative fluorescence unit.

Figure 3

Effect of treatment with CBE (100 µg/ml), chlorogenic acid (29.62 µg/ml), caffeine (5.11 µg/ml), caffeic acid (3.16 µg/ml), ferulic acid (1.54 µg/ml) and acarbose (50 µM) for 4 h at 37˚C, on sucrase enzyme activity. Values are presented as the mean ± the standard error of the mean (n=6). ^*P<0.05, ^**P<0.01, ^***P<0.001 vs. control. CBE, Coffea arabica bean extract.

Figure 4

Effect of treatment with CBE (100 µg/ml), chlorogenic acid (29.62 µg/ml), caffeine (5.11 µg/ml), caffeic acid (3.16 µg/ml), ferulic acid (1.54 µg/ml), phlorizin (0.5 mM) and phloretin (1 mM), for 4 h at 37˚C, on glucose transporters membrane protein expression. (A) Representative blot of SGLT1 membrane protein expression, (B) semi-quantification of relative SGLT1/ALP protein expression in each fraction, (C) representative blot of GLUT2 membrane protein expression and (D) semi-quantification of relative GLUT2/Na+K+ATPase protein expression in each fraction. Values are presented as the mean ± the standard error of the mean (n=3). ^*P<0.05, ^**P<0.01 vs. control. CBE, Coffea arabica bean extract; SGLT1, sodium glucose co-transporter 1; GLUT2 glucose transporter 2; ALP, alkaline phosphatase.

Figure 5

Effect of treatment with CBE (100 µg/ml), chlorogenic acid (29.62 µg/ml), caffeine (5.11 µg/ml), caffeic acid (3.16 µg/ml), ferulic acid (1.54 µg/ml), phlorizin (0.5 mM) and phloretin (1 mM), for 4 h at 37˚C on AMPK protein expression. (A) Representative blot of AMPK membrane protein expression and (B) semi-quantification of relative AMPK/β-actin protein expression in each fraction. Values are presented as the mean ± the standard error of the mean (n=3). ^*P<0.05 vs. control. CBE, Coffea arabica bean extract; p-, phospho; AMPK, 5-AMP-activated protein kinase.

Figure 6

mRNA expression levels of glucose transporter regulator genes, including SREBP-1c and HNF1α, in Caco-2 cells treated with CBE (100 µg/ml) and its constituents at a molar ratio concentration, including chlorogenic acid (29.62 µg/ml), caffeine (5.11 µg/ml), caffeic acid (3.16 µg/ml), ferulic acid (1.54 µg/ml), phlorizin (0.5 mM) or phloretin (1 mM), for 4 h at 37˚C. Values are presented as the mean ± the standard error of the mean (n=5). ^*P<0.05 vs. control. CBE, Coffea arabica bean extract; SREBP-1c, sterol regulatory element binding protein-1c; HNF1α, hepatocyte nuclear factor 1α.

Figure 7

Inhibitory effects of CBE on glucose transport and its mechanisms. CBE, Coffea arabica bean extract. SGLT1, sodium glucose co-transporter 1; GLUT2 glucose transporter 2; HNF1α, hepatocyte nuclear factor 1α;AMPK, 5-AMP-activated protein kinase.