Keiko Kokoroishi1, Ayumu Nakashima2,3, Shigehiro Doi4,5, Toshinori Ueno1, Toshiki Doi1, Yukio Yokoyama1, Kiyomasa Honda6, Masami Kanawa7, Yukio Kato6, Nobuoki Kohno8, Takao Masaki1. 1. Department of Nephrology, Hiroshima University Hospital, Hiroshima, Japan. 2. Department of Nephrology, Hiroshima University Hospital, Hiroshima, Japan. ayumu@hiroshima-u.ac.jp. 3. Department of Regeneration and Medicine, Medical Center for Translational and Clinical Research, Hiroshima University Hospital, 1-2-3 Kasumi Minami-Ku, Hiroshima, 734-8553, Japan. ayumu@hiroshima-u.ac.jp. 4. Department of Nephrology, Hiroshima University Hospital, Hiroshima, Japan. sdoi@hiroshima-u.ac.jp. 5. Department of Blood Purification, Hiroshima University Hospital, 1-2-3 Kasumi Minami-Ku, Hiroshima, 734-8553, Japan. sdoi@hiroshima-u.ac.jp. 6. Department of Dental and Medical Biochemistry, Graduate School of Biomedical and Health Sciences, Hiroshima University, Hiroshima, Japan. 7. Natural Science Center for Basic Research and Development, Hiroshima University, Hiroshima, Japan. 8. Department of Molecular and Internal Medicine, Graduate School of Biomedical and Health Sciences, Hiroshima University, Hiroshima, Japan.
Abstract
BACKGROUND: High glucose (HG) induces production of transforming growth factor-beta1 (TGF-β1), but the mechanism remains elusive. The aim of this study was to determine the gene(s) involved in HG-induced TGF-β1 production in human peritoneal mesothelial cells (HPMCs). METHODS: Microarray analysis was performed following a 3-h preincubation of HPMCs in 4 or 0.1% glucose medium. Transcriptional genes were selected using Gene Ontology analysis for biological processes, including regulation of transcription and DNA-dependent. The effects of small interfering RNA (siRNA) treatments on the up-regulation of TGF-β1 mRNA were assessed by quantitative real-time polymerase chain reaction (qPCR). Finally, enzyme-linked immunosorbent assay (ELISA) was performed to determine which gene(s) contribute to the production of TGF-β1 protein in the medium. RESULTS: Microarray analysis revealed that the expression of 51 genes increased by more than 3-fold. Gene ontology analysis identified 13 genes for further study. qPCR confirmed mRNA amplification for 9 of the 13 genes. Furthermore, HG-induced up-regulation of TGF-β1 mRNA was attenuated by the siRNA of 4 genes: MDS1 and EVI1 complex locus (MECOM), FBJ murine osteosarcoma viral oncogene homolog B (FOSB), FBJ murine osteosarcoma viral oncogene homolog (FOS) and activating transcription factor 3 (ATF3). ELISA showed that siRNA treatment of FOS, but not MECOM, FOSB or ATF3, suppressed the increase of TGF-β1 protein in the medium. CONCLUSIONS: FOS is a downstream effector of HG stimulation in HPMCs that contributes to TGF-β1 production, suggesting that blocking FOS expression may be a therapeutic target for peritoneal fibrosis.
BACKGROUND: High glucose (HG) induces production of transforming growth factor-beta1 (TGF-β1), but the mechanism remains elusive. The aim of this study was to determine the gene(s) involved in HG-induced TGF-β1 production in human peritoneal mesothelial cells (HPMCs). METHODS: Microarray analysis was performed following a 3-h preincubation of HPMCs in 4 or 0.1% glucose medium. Transcriptional genes were selected using Gene Ontology analysis for biological processes, including regulation of transcription and DNA-dependent. The effects of small interfering RNA (siRNA) treatments on the up-regulation of TGF-β1 mRNA were assessed by quantitative real-time polymerase chain reaction (qPCR). Finally, enzyme-linked immunosorbent assay (ELISA) was performed to determine which gene(s) contribute to the production of TGF-β1 protein in the medium. RESULTS: Microarray analysis revealed that the expression of 51 genes increased by more than 3-fold. Gene ontology analysis identified 13 genes for further study. qPCR confirmed mRNA amplification for 9 of the 13 genes. Furthermore, HG-induced up-regulation of TGF-β1 mRNA was attenuated by the siRNA of 4 genes: MDS1 and EVI1 complex locus (MECOM), FBJ murineosteosarcoma viral oncogene homolog B (FOSB), FBJ murineosteosarcoma viral oncogene homolog (FOS) and activating transcription factor 3 (ATF3). ELISA showed that siRNA treatment of FOS, but not MECOM, FOSB or ATF3, suppressed the increase of TGF-β1 protein in the medium. CONCLUSIONS:FOS is a downstream effector of HG stimulation in HPMCs that contributes to TGF-β1 production, suggesting that blocking FOS expression may be a therapeutic target for peritoneal fibrosis.
Entities:
Keywords:
FOS; High glucose; Human peritoneal mesothelial cells; Microarray; TGF-β1
Authors: Cora Weigert; Ulrich Sauer; Katrin Brodbeck; Andreas Pfeiffer; Hans U Häring; Erwin D Schleicher Journal: J Am Soc Nephrol Date: 2000-11 Impact factor: 10.121
Authors: Felipe Simon; Pablo Tapia; Ricardo Armisen; Cesar Echeverria; Sebastian Gatica; Alejandro Vallejos; Alejandro Pacheco; Maria E Sanhueza; Miriam Alvo; Erico Segovia; Rubén Torres Journal: Front Physiol Date: 2017-06-13 Impact factor: 4.566