Effects of consumption of whole grape powder on basal NF-κB signaling and inflammatory cytokine secretion in a mouse model of inflammation

Abstract

Dietary consumption of polyphenol-rich fruits, such as grapes, may reduce inflammation and potentially prevent diseases linked to inflammation. Here, we used a genetically engineered murine model to measure Nuclear Factor kappa-light-chain-enhancer of activated B cells (NF-κB) activity and pro-inflammatory cytokine secretion to test the hypothesis that oral consumption of whole grape formulation reduces inflammatory signaling in the body. NF-κB luciferase reporter mice were divided into two groups, one which was fed an experimental diet formulated with 4% (w/w) whole grape powder (WGP) or another which was fed a control diet formulated with 3.6% glucose/fructose (w/w) combination. Simulated inflammation was induced in the mice by intraperitoneal injection of lipopolysaccharide (LPS). In vivo imaging was used to determine the effect of each diet on NF-κB activity. We found that there were no significant differences in weight gain between the WGP and control diet groups. However, there was a statistically significant (p<0.0001) difference in the progression of basal levels of NF-κB signaling between mice fed on control or WGP diet. There were no significant differences in NF-κB reporter indices between WGP- and control-diet groups after either acute or repeated inflammatory challenge. However, terminal blood collection revealed significantly (p<0.01) lower serum concentrations of the inflammatory cytokines Interleukin-6 (IL-6) and Tumor Necrosis Factor alpha (TNFα) only among WGP diet mice subjected to acute inflammatory challenge. Overall, these data suggest that while diets supplemented with WGP may suppress steady-state low levels of inflammatory signaling, such a supplementation may not alleviate exogenously induced massive NF-κB activation.

Keywords: NF-kB; Whole grape powder; diet; in vivo; inflammation; mouse.

Figure 1

Bodyweight trend of mice on control or whole grape powder (WGP) supplemented diet. Mice were randomly grouped into either acute challenge or repeat challenge group, each of which was further sub-grouped into control or WGP diet. Weekly bodyweight measurements were taken. Plots represent average weight of animals (n=10 per group) versus time in weeks. Panel A is shows data plot for animals in the acute challenge group, while panel B shows plot for the repeat challenge group. WGP supplementation for 3 weeks did not affect bodyweight trend.

Figure 2

Effect of WGP supplementation on changes in basal NF-κB activity in reporter mice. Paired-sample plots for basal NF-κB reporter activity in mice fed control (Panel A, n=20) or WGP (Panel B, n=20) diet for 3 weeks. Changes in reporter activity over the 3 weeks for each of the animals in either control- or WGP-diet group are shown in panel C. Panel D shows luminescence images from mouse with no luciferin injection (left) and from a mouse in control diet group (at week 1 and week 3, middle panels), and from a mouse in WGP diet group (also at week 1 and week 3, right panels). The WGP group had significantly lower rate of increase in reporter activity over the 3 weeks period. ** statistically significant difference (p<0.0001).

Figure 3

Effect of WGP supplementation on changes in NF-κB reporter activity in mice subjected to acute inflammatory challenge. After 3 weeks on diet, mice were subjected to an acute inflammatory challenge. Changes in reporter activity were plotted as the difference between the pre-challenge (week 3 readings) and post-acute challenge readings. Box plots represent the differences in reporter activity for the whole body (panel A) and abdominal/peritoneal (panel B) regions of interest. There was no significant difference between control and WGP groups.

Figure 4

Effect of WGP supplementation on changes in NF-κB reporter activity in mice subjected to repeat inflammatory challenge (on days 1, 3, and 5). Changes in reporter activity were plotted as the difference between the pre-challenge (week 3 readings) and post- challenge readings on each day of challenge. Box plots represent the differences in reporter activity for the whole body (panel A) and abdominal/peritoneal (panel B) regions of interest. There were no significant differences between control and WGP groups for days 1 & 3 (p>0.06). Significance was reached (*, p=0.03) for day 5.

Figure 5

Effect of WGP supplementation on serum levels of the cytokines IL-6 and TNF-α. Terminal sera collected from mice on control or WGP diet were analyzed using ELISA for the levels of IL-6 and TNFα. IL-6 and TNFα data for mice in the acute challenge group are displayed in panels A and B, respectively. IL-6 and TNFα data for mice in the repeat challenge group are shown in panels C and D, respectively. ** statistically significant difference.