Fig 1.
(A) Effect of 24 h treatment with 5 mM acetate (ACET), butyrate (BUT), or propionate (PROP) on morphology of 3D murine duodenal enteroids as well as (B) EdU cellular proliferation marker (green), and F-actin distribution (red) compared to controls (CON).
Nuclei are stained in with DAPI (blue) and images are representative of several fields of view at 20X magnification.
Fig 2.
Mouse enteroids treated with single SCFAs alters gene expression after 24h.
Effect of 24 h treatment with acetate (ACET), butyrate (BUT), or propionate (PROP) at (A) 1 mM and (B) 5 mM. Gene expression of differentiation markers: stem cells (Lgr5), enterocyte alkaline phosphatase (Alpi), goblet cell mucin 2 (Muc2), enteroendocrine cell chromogranin A (Chga), Paneth cell lysozyme (Lyz), tight junction markers claudin-3 (Cldn3), occludin (Ocln), zonula-occludens 1 (Tjp1), Tuft cell marker Dclk1, anti-microbial peptide expression Reg3β, Reg3γ, and beta-defensin 1 (Defb1) were analyzed in murine 3D duodenal enteroids. Treatments are compared to controls (CON) which are set to 1.0 and superscripts (a,b,c,d) indicate statistical significance between treatments at P < 0.05, n = 4.
Fig 3.
Effect of 24 h treatment with 10 mM acetate (ACET), butyrate (BUT), or propionate (PROP) on morphology of 3D human duodenal enteroids (A) and 3D human small intestinal enteroid cell proliferation (EdU staining, green) compared to controls (CON). Nuclei are stained with DAPI (blue) and images are representative of several fields of view at 20X magnification (B). Treatments are compared to controls (CON) which are set to 1.0 and superscripts (a,b,c,d) indicate statistical significance between treatments at P < 0.05, n = 6.
Fig 4.
Effect of 24 h treatment with acetate (ACET), butyrate (BUT), or propionate (PROP) at (A) 1 mM and (B) 10 mM.
Gene expression of differentiation markers: stem cells (Lgr5), enterocyte alkaline phosphatase (Alpi), goblet cell mucin 2 (Muc2), enteroendocrine cell chromogranin A (Chga), Paneth cell lysozyme (Lyz), tight junction markers claudin-3 (Cldn3), occludin (Ocln), zonula-occludens 1 (Tjp1), Tuft cell marker Dclk1, anti-microbial peptide expression Reg3β, Reg3γ, and beta-defensin 1 (Defb1) were analyzed in human 3D duodenal enteroids. Treatments are compared to controls (CON) which are set to 1.0 and superscripts (a,b,c,d) indicate statistical significance between treatments at P < 0.05, n = 4.
Fig 5.
Effect of 24 h treatment with 10 mM acetate (ACET), butyrate (BUT), or propionate (PROP) on 3D (a) and 2D (b) human small intestinal enteroid cell proliferation (EdU staining, green) compared to controls (CON).
Nuclei are stained with DAPI (blue) and images are representative of several fields of view at 20X magnification. Treatments are compared to controls (CON) which are set to 1.0 and superscripts (a,b,c,d) indicate statistical significance between treatments at P < 0.05, n = 6.
Fig 6.
Effect of 24 h treatment with acetate (ACET), butyrate (BUT), or propionate (PROP) at 10 mM.
Gene expression of differentiation markers: stem cells (Lgr5), enterocyte alkaline phosphatase (Alpi), goblet cell mucin 2 (Muc2), enteroendocrine cell chromogranin A (Chga), Paneth cell lysozyme (Lyz), tight junction markers claudin-3 (Cldn3), occludin (Ocln), zonula-occludens 1 (Tjp1), Tuft cell marker Dclk1, anti-microbial peptide expression Reg3β, Reg3γ, and beta-defensin 1 (Defb1) were analyzed in human 2D duodenal enteroids. Treatments are compared to controls (CON) which are set to 1.0 and superscripts (a,b,c,d) indicate statistical significance between treatments at P < 0.05, n = 4.
Table 1.
Protein expression of neuroendocrine hormones and cytokines in human 2D duodenal enteroids treated with short chain fatty acids.
Fig 7.
(A) Alcian blue/Hematoxylin staining 2D enteroids treated with 10 mM acetate (ACET), butyrate (BUT), and propionate (PROP) compared to controls (CON). (B) Zonula occludens-1 (ZO-1) staining (red) and (C) Claudin-3 (CLDN3) staining (red). Nuclei are stained with DAPI (blue). (D) Western blot data for Chromogranin-A (CHGA), Zonula-occludens-1 (ZO-1), occludin (OCLN), and Trefoil Factor 3 (TFF3). Samples were compared to β-actin as a housekeeping protein. Confocal images are representative of several fields of view at 40X magnification. Treatments are compared to controls (CON) which are set to 1.0 and superscripts (a,b,c,d) indicate statistical significance between treatments at P < 0.05, n = 4.
Fig 8.
Effect of 24 h treatment with 10 mM acetate (ACET), butyrate (BUT), or propionate (PROP) on (A) epithelial barrier function of human duodenal enteroids as measured by TER compared to controls (CON). Values are represented as relative change in TER at time 0 and time 24 h after treatment. (B) Raw TER values at 0 h and 24 h post-treatment. Treatments are compared to controls (CON) which are set to 1.0 and superscripts (a,b,c,d) indicate statistical significance between treatments at P < 0.05, n = 4.