Figure 1.
Lanthionine synthetase C-like 2 (LANCL2) and NSC61610 in silico.
(A) The homology model of human LANCL2 is shown in Cartoon representation with coloring according to secondary structure. Purple: alpha helix; Blue: other helix; Yellow: bridge_beta; Cyan: turn; Green: coil. (B) 2-D structure of NSC61610. (C) Representative binding mode of the most stable docked orientation of NSC61610 with LANCL2. The LANCL2 model is shown in ribbon mode. NSC61610 pose generated by AutoDock Vina is colored in cyan and the one generated by AutoDock is colored in orange. Selected residues of LANCL2 (blue) are depicted by stick-and-ball models and colored by atom types. Amino acid residues surrounding NSC61610 are labeled. (D) Representative binding modes of the docked orientation of abscisic acid (ABA) and NSC61610 with LANCL2. LANCL2 is shown in a ribbon mode. NSC61610 (orange) and ABA (magenta) are shown in stick-and-ball model. Selected residues of LANCL2 surrounding both NSC61610 and ABA are depicted by stick-and-ball model and labeled. The images were rendered in Visual Molecular Dynamics (VMD).
Figure 2.
Effect of LANCL2 disruption and cAMP inhibition on PPAR γ activation in RAW 264.7 macrophages.
Cells were cotransfected with a pTK.PPRE3x luciferase reporter plasmid driven by the PPRE-containing Acyl-CoA oxidase promoter with or without LANCL2 siRNA. Then, cells were treated with vehicle (DMSO) or NSC61610 (2.5 µM), the adenylate cyclase-specific inhibitor 2′5′-dideoxyadenosine (10 µM). Luciferase activity was normalized to pRL activity in the cell extracts and relative luciferase activity was calculated a ratio of the activity in the treatment wells to control wells. Data are represented as mean ± standard error. Points with an asterisk indicate that a treatment is significantly different from its control (P<0.05).
Figure 3.
Oral treatment with NSC61610 ameliorates experimental inflammatory bowel disease.
Mice were challenged with 2.5% dextran sodium sulfate in the drinking water for 7 days. Disease activity index (DAI), a composite score reflecting clinical signs of the disease (i.e. perianal soiling, rectal bleeding, diarrhea and piloerection), was assessed daily. Panel A illustrates the effect of NSC61610 on disease severity in mice with colitis. Panels B–D illustrate the effect of NSC61610 on macroscopic inflammatory lesions in the colon (B), spleen (C), and mesenteric lymph nodes (MLN) (D). Data are represented as mean ± standard error (n = 10). In figure A, data points with asterisks are significantly different from control and data points with two asterisks are significantly different from those with one asterisk (P<0.05). In figure B–D, bars with an asterisk indicate that a treatment is significantly different from its control (P<0.05).
Figure 4.
Oral treatment with NSC61610 ameliorates inflammatory lesions in mice with inflammatory bowel disease.
Mice were challenged with 2.5% dextran sodium sulfate in the drinking water for 7 days. Representative photomicrographs from the control (A–B) and NSC61610 treatment (C–E) groups are illustrated. Colonic specimens underwent blinded histological examination and were scored based on epithelial erosion (F), mucosal wall thickening (G), and leukocyte infiltration (H). Data are represented as mean ± standard error (n = 10). Bars with an asterisk indicate that a treatment is significantly different from its control (P<0.05).
Figure 5.
Modulation of colonic gene expression by oral treatment with NSC61610.
Mice were challenged with 2.5% dextran sodium sulfate in the drinking water for 7 days. Colonic mRNA expression of peroxisome proliferator-activated receptor γ (PPAR γ) (A), monocyte chemoattractant protein-1 (MCP-1) (B), and interleukin-6 (IL-6) (C) were assessed by quantitative real-time RT-PCR. Data are represented as mean ± standard error (n = 10). Bars with an asterisk indicate that a treatment is significantly different from its control (P<0.05).
Figure 6.
Oral treatment with NSC61610 on the distribution of immune cell subsets in colonic lamina propria and spleen.
Colonic lamina propria lymphocytes (LPL, A, B, and E) and spleen (C and D), were immunophenotyped to identify regulatory T cells (Treg) and CD4+IL-10+ T cell subsets through flow cytometry. Data are represented as mean ± standard error (n = 10). Bars with an asterisk indicate that a treatment is significantly different from its control (P<0.05).
Figure 7.
Oral treatment with NSC61610 on the distribution of macrophages in colonic lamina propria.
Colonic lamina propria lymphocytes (LPL) were immunophenotyped to identify F4/80+CD11b+ macrophage subsets through flow cytometry. Data are represented as mean ± standard error (n = 10). Bars with an asterisk indicate that a treatment is significantly different from its control (P<0.05).
Figure 8.
Effect of tissue-specific PPAR γ deletion and oral NSC61610 treatment in experimental inflammatory bowel disease.
Mice were challenged with 2.5% dextran sodium sulfate in the drinking water. Panel A illustrates the effect of oral NSC61610 treatment on disease severity. Means within time points with different letterssuperscripts are significant different (P<0.05). Panels B–D illustrate the effect of oral NSC61610 on macroscopic lesions in the colon (B), spleen (C), and mesenteric lymph nodes (MLN) (D). Data are represented as mean ± standard error (n = 10). Bars with an asterisk indicate that a treatment is significantly different from its control (P<0.05).