Upregulation of REG IV gene in human intestinal epithelial cells by lipopolysaccharide via downregulation of microRNA-24.

The pathophysiology of inflammatory bowel diseases (IBD) reflects a balance between mucosal injury and reparative mechanisms. Some regenerating gene (Reg) family members (REG Iα, REG Iß and REG IV) are expressed in Crohn's disease (CD) and ulcerative colitis (UC) and involved as proliferative mucosal factors in IBD. We revealed that REG Iα and REG Iß were induced in cell culture system by IL-6/IL-22. Although REG IV was upregulated in IBD biopsy samples, the upregulation of REG IV was not at all induced in cell culture by autoimmune-related cytokines such as IL-6, IL-22 and TNFα. Here, we analysed REG IV expression in LS-174 T and HT-29 human intestinal epithelial cells by real-time RT-PCR and elisa. REG IV expression was induced by lipopolysaccharide (LPS). However, LPS did not activate REG IV promoter activity. As the LPS-induced upregulation of REG IV was considered to be regulated post-transcriptionally, we searched targeted microRNA (miR), which revealed that REG IV mRNA has a potential target sequence for miR-24. We measured the miR-24 level of LPS-treated cells and found that the level was significantly lower. The LPS-induced increase of REG IV mRNA was abolished by the introduction of miR-24 mimic but not by non-specific control RNA.

Expression of human REG family genes in inflammatory bowel disease and their molecular mechanism.

The pathophysiology of inflammatory bowel disease (IBD) reflects a balance between mucosal injury and reparative mechanisms. Some regenerating gene (Reg) family members have been reported to be expressed in Crohn's disease (CD) and ulcerative colitis (UC) and to be involved as proliferative mucosal factors in IBD. However, expression of all the REG family genes in IBD is still unclear. Here, we analyzed expression of all the REG family genes (REGIα, REGIß, REG III, HIP/PAP, and REG IV) in biopsy specimens of UC and CD by real-time RT-PCR. REG Iα, REG Iß, and REG IV genes were overexpressed in CD samples. REG IV gene was also overexpressed in UC samples. We further analyzed the expression mechanisms of REG Iα, REG Iß, and REG IV genes in LS-174T and HT-29 human colonic epithelial cells. The expression of REG Iα was significantly induced by IL-6 or IL-22, and REG Iß was induced by IL-22. Deletion analyses revealed that three regions (- 220~- 211, - 179~- 156, and - 146~- 130) in REG Iα and the region (- 274~- 260) in REG Iß promoter were responsible for the activation by IL-22/IL-6. The promoters contain consensus transcription factor binding sequences for MZF1, RTEF1/TEAD4, and STAT3 in REG Iα, and HLTF/FOXN2F in REG Iß, respectively. The introduction of siRNA for MZF1, RTEF1/TEAD4, STAT3, and HLTF/FOXN2F abolished the transcription of REG Iα and REG Iß. The gene activation mechanisms of REG Iα/REG Iß may play a role in colon mucosal regeneration in IBD.

Expression of REG family genes in human inflammatory bowel diseases and its regulation.

The pathophysiology of inflammatory bowel disease (IBD) reflects a balance between mucosal injury and reparative mechanisms. Some regenerating gene (Reg) family members have been reported to be expressed in Crohn's disease (CD) and ulcerative colitis (UC) and to be involved as proliferative mucosal factors in IBD. However, expression of all REG family genes in IBD is still unclear. Here, we analyzed expression of all REG family genes (REG Iα, REG Iß, REG III, HIP/PAP, and REG IV) in biopsy specimens of UC and CD by real-time RT-PCR. REG Iα, REG Iß, and REG IV genes were overexpressed in CD samples. REG IV gene was also overexpressed in UC samples. We further analyzed the expression mechanisms of REG Iα, REG Iß, and REG IV genes in human colon cells. The expression of REG Iα was significantly induced by IL-6 or IL-22, and REG Iß was induced by IL-22. Deletion analyses revealed that three regions (- 220 to - 211, - 179 to - 156, and - 146 to - 130) in REG Iα and the region (- 274 to- 260) in REG Iß promoter were responsible for the activation by IL-22/IL-6. The promoters contain consensus transcription factor binding sequences for MZF1, RTEF1/TEAD4, and STAT3 in REG Iα, and HLTF/FOXN2F in REG Iß, respectively. The introduction of siRNAs for MZF1, RTEF1/TEAD4, STAT3, and HLTF/FOXN2F abolished the transcription of REG Iα and REG Iß. The gene activation mechanisms of REG Iα/REG Iß may play a role in colon mucosal regeneration in IBD.

Synergistic activations of REG I α and REG I ß promoters by IL-6 and Glucocorticoids through JAK/STAT pathway in human pancreatic ß cells.

Reg (Regenerating gene) gene was originally isolated from rat regenerating islets and its encoding protein was revealed as an autocrine/paracrine growth factor for ß cells. Rat Reg gene is activated in inflammatory conditions for ß cell regeneration. In human, although five functional REG family genes (REG Iα, REG Iß, REG III, HIP/PAP, and REG IV) were isolated, their expressions in ß cells under inflammatory conditions remained unclear. In this study, we found that combined addition of IL-6 and dexamethasone (Dx) induced REG Iα and REG Iß expression in human 1.1B4 ß cells. Promoter assay revealed that a signal transducer and activator of transcription- (STAT-) binding site in each promoter of REG Iα (TGCCGGGAA) and REG Iß (TGCCAGGAA) was essential for the IL-6+Dx-induced promoter activation. A Janus kinase 2 (JAK2) inhibitor significantly inhibited the IL-6+Dx-induced REG Iα and REG Iß transcription. Electrophoretic mobility shift assay and chromatin immunoprecipitation revealed that IL-6+Dx stimulation increased STAT3 binding to the REG Iα promoter. Furthermore, small interfering RNA-mediated targeting of STAT3 blocked the IL-6+Dx-induced expression of REG Iα and REG Iß. These results indicate that the expression of REG Iα and REG Iß should be upregulated in human ß cells under inflammatory conditions through the JAK/STAT pathway.