N-butyric acid induces EBV virus production.

<p>Flow cytometric analysis of GFP fluorescence in Raji cells upon superinfection with EBV-GFP recombinant virus produced by EBV-GFP carrying AGS epithelial cells, upon treatment with n-butyric acid as stated in the M&M.</p

Short Chain Fatty Acids (SCFA) Reprogram Gene Expression in Human Malignant Epithelial and Lymphoid Cells

<div><p>The effect of short chain fatty acids (SCFAs) on gene expression in human, malignant cell lines was investigated, with a focus on signaling pathways. The commensal microbial flora produce high levels of SCFAs with established physiologic effects in humans. The most abundant SCFA metabolite in the human microflora is n-butyric acid. It is well known to activate endogenous latent Epstein-Barr virus (EBV), that was used as a reference read out system and extended to EBV+ epithelial cancer cell lines. N-butyric acid and its salt induced inflammatory and apoptotic responses in tumor cells of epithelial and lymphoid origin. Epithelial cell migration was inhibited. The n-butyric gene activation was reduced by knock-down of the cell membrane transporters MCT-1 and -4 by siRNA. N-butyric acid show biologically significant effects on several important cellular functions, also with relevance for tumor cell phenotype.</p></div

N-butyric acid induces proinflammatory cytokine expression.

<p>A. qRT-PCR analysis of Il-6 and IL-8 expression after overnight exposure to 10mM n-butyric acid. Values are adjusted to the EF1α gene expression. B. ELISA analysis of IL-8 secretion from C666-1 cells after 48hr of exposure to 10mM n-butyric acid.</p

N-butyric acid inhibits migration of C666-1 NPC cells.

<p>Evaluation of the gap distance in migration assay of C666-1 NPC cells upon treatment as indicated.</p

Analysis of short chain fatty acids induced BZLF1 gene expression in C666-1 NPC cells.

<p>RT-PCR analysis upon overnight treatment with 10mM of short chain fatty acids as indicated. A. BZLF1 expression. B. BART A expression. C. GAPDH expression. Cells were treated with 10mM of SCFAs as indicated or were left untreated. D. Digital analysis of data from panels A-C, demonstrating BZLF1 expression in relation to GAPDH.</p

N-butyric acid induces PARP cleavage.

<p>Western blot analysis of cells treated with butyrate or TPA for 24hr, A-C) Raji cells, D-F) C666-1 cells. A,D) PonceauS staining. C,E) Actin. B,F) anti PARP Ab.</p

A comparison of the differentially activated signaling pathways in LMP2A positive cells.

<p>A comparison of the differentially activated signaling pathways in LMP2A positive cells.</p

N-butyric acid induces NFkappaB signaling.

<p>Western blot analysis of phospho-IkappaB and NFkappaB expression in treated C666-1 (A) and Raji cells (B), adjusted to IkappaB and GAPDH expression, respectively.</p

N-butyric acid enters cells through MCT1 and 4 transporters.

<p>A. RT-PCR analysis of MCT1 and 4 expression in C666-1 NPC cells upon n-butyric acid treatment during 24hr. B. RT-PCR analysis of MCT expression in Raji B cells upon different treatments during 24hr. Lanes represent the same as in A. C. Graph represents MCT1 and 4 gene expression in non treated cells, measured as pixel density and adjusted to GAPDH expression. D. The left panel. Expression of BZLF1, the viral lytic message, and cellular MCT transporters in C666-1 cells, adjusted to BART A and GAPDH, respectively, is decreased after knock-down of MCT, followed by n-butyric acid treatment. The right panel. qRT-PCR analysis of IL-8 expression in these cells. Pixel density values are compared to those after HCl treatment.</p

Genes expression of which changed in HONE1 cells treated with 10mM Butyric Acid for 24hr, were enriched for four different signaling-related GO terms.

<p>Genes expression of which changed in HONE1 cells treated with 10mM Butyric Acid for 24hr, were enriched for four different signaling-related GO terms.</p