Yan Chen1, Zheng-Yang Li, Yan Yang, Hong-Jie Zhang. 1. Department of Gastroenterology, First Affiliated Hospital of Nanjing Medical University, Nanjing 210029, Jiangsu Province, China.
Abstract
AIM: To investigate whether uncoupling protein 2 (UCP2) affects oleic acid-induced secretion of glucagon-like peptide-1 (GLP-1) in L-cells. METHODS: mRNA and protein expression of UCP2 were analyzed in human NCI-H716 cells, which serve as a model for enteroendocrine L-cells, by quantitative reverse transcription-polymerase chain reaction and Western blotting before and after treatment with oleic acid. Localization of UCP2 and GLP-1 in NCI-H716 cells was assessed by immunofluorescence labeling. NCI-H716 cells were transiently transfected with a small interfering RNA (siRNA) that targets UCP2 (siUCP2) or with a non-specific siRNA using Lipofectamine 2000. The concentrations of bioactive GLP-1 in the medium were measured by enzyme linked immunosorbent assay. RESULTS: Both GLP-1 and UCP2 granules were expressed mainly in the cytoplasm of NCI-H716 cells. NCI-H716 cells that secreted GLP-1 also expressed UCP2. Time-course experiments revealed that release of GLP-1 from NCI-H716 cells into the medium reached a maximum at 120 min and remained stable until at least 180 min after treatment with oleic acid (the level of GLP-1 increased about 2.3-fold as compared with the level of GLP-1 in the control cells, P < 0.05). In an experiment to determine dose dependence, stimulation of NCI-H716 cells with ≤ 8 mmol oleic acid led to a concentration-dependent release of GLP-1 into the medium; 10 mmol oleic acid diminished the release of GLP-1. Furthermore, GLP-1 secretion induced by oleic acid from NCI-H716 cells that were transfected with siUCP2 decreased to 41.8%, as compared with NCI-H716 cells that were transfected with a non-specific siRNA (P < 0.01). CONCLUSION: UCP2 affected GLP-1 secretion induced by oleic acid. UCP2 plays an important role in L-cell secretion that is induced by free fatty acids.
AIM: To investigate whether uncoupling protein 2 (UCP2) affects oleic acid-induced secretion of glucagon-like peptide-1 (GLP-1) in L-cells. METHODS: mRNA and protein expression of UCP2 were analyzed in human NCI-H716 cells, which serve as a model for enteroendocrine L-cells, by quantitative reverse transcription-polymerase chain reaction and Western blotting before and after treatment with oleic acid. Localization of UCP2 and GLP-1 in NCI-H716 cells was assessed by immunofluorescence labeling. NCI-H716 cells were transiently transfected with a small interfering RNA (siRNA) that targets UCP2 (siUCP2) or with a non-specific siRNA using Lipofectamine 2000. The concentrations of bioactive GLP-1 in the medium were measured by enzyme linked immunosorbent assay. RESULTS: Both GLP-1 and UCP2 granules were expressed mainly in the cytoplasm of NCI-H716 cells. NCI-H716 cells that secreted GLP-1 also expressed UCP2. Time-course experiments revealed that release of GLP-1 from NCI-H716 cells into the medium reached a maximum at 120 min and remained stable until at least 180 min after treatment with oleic acid (the level of GLP-1 increased about 2.3-fold as compared with the level of GLP-1 in the control cells, P < 0.05). In an experiment to determine dose dependence, stimulation of NCI-H716 cells with ≤ 8 mmol oleic acid led to a concentration-dependent release of GLP-1 into the medium; 10 mmol oleic acid diminished the release of GLP-1. Furthermore, GLP-1 secretion induced by oleic acid from NCI-H716 cells that were transfected with siUCP2 decreased to 41.8%, as compared with NCI-H716 cells that were transfected with a non-specific siRNA (P < 0.01). CONCLUSION:UCP2 affected GLP-1 secretion induced by oleic acid. UCP2 plays an important role in L-cell secretion that is induced by free fatty acids.