Table 1.
List of PCR primers.
Fig 1.
Effects of butyrate and LPS on intestinal barrier integrity measured by TEER and paracellular permeability in IPEC-J2 cells.
(A) Cells were challenged with LPS on day 9 post-differentiation; TEER was measured at 0, 12, and 24 h after LPS challenge, respectively. (B) Paracellular permeability was determined by measuring FITC-dextran flux at 24 h after LPS challenge. Data were analyzed by one-way ANOVA with Tukey multiple comparison test. Values are means ± SE, n = 4. Different superscript letters on bars (a, b, c, d, e) indicate significant mean differences, P < 0.05.
Fig 2.
Effect of butyrate and LPS on mRNA expression of tight junction proteins in IPEC-J2 cells.
Real time PCR was performed to determine mRNA expression of (A) claudin-1, (B) claudin-3, (C) claudin-4, (D) occludin and (E) ZO-1 at 4 and 8 h after LPS challenge, respectively. Data were analyzed by one-way ANOVA with Tukey multiple comparison test. Values are means ± SE, n = 6. Different superscript letters on bars (a, b, c) indicate significant differences, P < 0.05.
Fig 3.
Effect of butyrate and LPS on abundance of tight junction proteins in IPEC-J2 cells.
Cell lysates were isolated after 4 and 8 h of LPS challenge, and subjected to immunoblotting analysis. Data were analyzed by one-way ANOVA with Tukey multiple comparison test. Values are means ± SE, n = 4. Different superscript letters (a, b, c) on bars indicate significant mean differences, P < 0.05.
Fig 4.
Effect of butyrate and LPS on mRNA expression of inflammatory related genes and secretion of IL-8 in IPEC-J2 cells.
(A) mRNA expression of inflammation related genes after 8 h LPS challenge. (B) Level of secretory IL-8 in cell culture media after 8 h of LPS. Data were analyzed by one-way ANOVA with Tukey multiple comparison test. Values are means ± SE, n = 6. Different superscript letters (a, b, c, d) on bars indicate significant mean differences, P < 0.05.
Fig 5.
Effect of butyrate and LPS treatment on activation of Akt in IPEC-J2 cells.
(A) Representative immunoblotting analysis of Akt after 4 and 8 h of LPS stimulation. (B) Quantification analysis of Akt abundance in IPEC-J2 cells. Data were analyzed by one-way ANOVA with Tukey multiple comparison test. Values are means ± SE, n = 4. Different superscript letters (a, b, c) on bars indicate significant mean differences, P < 0.05.
Fig 6.
Effect of butyrate and LPS challenge on phosphorylation of 4E-BP-1 in IPEC-J2 cells.
(A) Representative immunoblotting analysis of 4E-BP-1 after 4 and 8 h of LPS stimulation. (B) Quantification analysis of Akt abundance in IPEC-J2 cells. Data were analyzed by one-way ANOVA with Tukey multiple comparison test. Values are means ± SE, n = 4. Different superscript letters on bars (a, b, c) indicate significant mean differences, P < 0.05.
Fig 7.
Effect of butyrate and LPS on intracellular ATP level and AMPK activation in IPEC-J2 cells.
Cell lysates were isolated after 8 h of LPS challenge, and subjected to ATP determination assay and immunoblotting analysis. (A) Intracellular ATP level was measured by ATP determination kit and normalized to total protein concentration. (B) Representative immunoblotting and quantification analyses of AMPK abundance in IPEC-J2 cells. Data were analyzed by one-way ANOVA with Tukey multiple comparison test. Values are means ± SE, n = 4. Different superscript letters on bars (a, b, c) indicate significant mean differences, P < 0.05.