Assessment of chitosan-affected metabolic response by peroxisome proliferator-activated receptor bioluminescent imaging-guided transcriptomic analysis.

Construction and optimization of PPRE reporter constructs.(A) The schematic diagram of PPRE reporter constructs. Two PPRE oligonucleotides were annealed and ligated to form various tandem repeats of PPRE. The resulting products were analyzed by 8% polyacrylamide gels (left panel). Eight reporter constructs containing various numbers of PPREs were shown on the right. (B) Effect of rosiglitazone on the inducibility of PPRE reporter constructs. HepG2 cells were transiently transfected with PPRE constructs and pcDNA3.1/lacZ DNA, and treated without or with 0.5 µM rosiglitazone. Luciferase and β-galactosidase activities were determined 24 hours later. Luciferase activities are expressed as induction fold, which is presented as comparison with RLU related to untreated cells. β-Galactosidase activities are expressed as OD420. Values are mean ± standard error of three independent assays. **p<0.01, ***p<0.001, compared with untreated cells. (C) In vitro imaging. HepG2 cells were transiently transfected with PPRE constructs containing 5 tandem repeats of PPRE and treated without or with 0.5 µM rosiglitazone. Luciferase activity was imaged at 24 h by IVIS system. The color overlay on the image represents the photon/sec emitted from the cells, as indicated by the color scale. Quantification of photon emission from the cells was shown at the bottom. Values are mean ± standard error of three independent assays. ***p<0.001, compared with mock. Photos are representative images.

PPAR-dependent bioluminescence in living mice and individual organs.(A) In vivo imaging. Transgenic mice were orally administered saline (mock) or 50 mg/kg rosiglitazone and imaged 6 hours later. The color overlay on the image represents the photon/sec emitted from the animals, as indicated by the color scales. Quantification of photon emission from the mice was shown at the bottom. Values are mean ± standard error (n=6 per group). *p<0.05, **p<0.01, compared with mock. (B) Ex vivo imaging. Transgenic mice were orally administered saline (mock) or 50 mg/kg rosiglitazone. Six hours later, mice were sacrificed and organs were subjected to image. Quantification of photon emission from the organs was shown at the bottom. Values are mean ± standard error (n=6 per group). Photos are representative images.

PPAR-dependent bioluminescence in living mice and individual organs after chitosan administration.(A) Time course. Transgenic mice were subcutaneously injected saline (mock) or chitosan, and images at indicated periods. Results are expressed as relative intensity, which is presented as comparison with the luminescent intensity relative to mock. Values are mean ± standard error (n=6 per group). (B) Ex vivo imaging and quantification of photon emission from individual organs. Transgenic mice were subcutaneously injected saline (mock) or chitosan, and sacrificed 3 days later for organ imaging. Values are mean ± standard error (n=6 per group). *p<0.05, ***p<0.001, compared with mock. The color overlay on the image represents the photon/sec emitted from the organs, as indicated by the color scale. Photos are representative images.