Yan Ding1,2, Lanlan Shan1, Wenqing Nai1, Xiaojun Lin3, Ling Zhou4, Xiaoying Dong4, Hongyuan Wu1, Min Xiao3, Xuejuan Zhou3, Linlin Wang4, Ting Li3, You Fu1, Yijun Lin1, Chunhong Jia3, Meng Dai1, Xiaochun Bai3,5. 1. From the Department of Health Management, Nanfang Hospital, Southern Medical University, Guangzhou, China. 2. Department of Endocrinology, Wuhan General Hospital of Guangzhou Command, Wuhan, Hubei Province, Southern Medical University, Guangzhou, China. 3. Department of Cell Biology, School of Basic Medical Sciences, Southern Medical University, Guangzhou, China. 4. Department of Oncology, Nanfang Hospital, Southern Medical University, Guangzhou, China. 5. State Key Laboratory of Organ Failure Research, Department of Cell Biology, School of Basic Medical Sciences, Southern Medical University, Guangzhou, China.
Abstract
BACKGROUND/AIMS: The mechanistic target of rapamycin (mTOR) signaling pathway is essential for angiogenesis and embryonic development. DEP domain-containing mTOR-interacting protein (DEPTOR) is an mTOR binding protein that functions to inhibit the mTOR pathway In vitro experiments suggest that DEPTOR is crucial for vascular endothelial cell (EC) activation and angiogenic responses. However, knowledge of the effects of DEPTOR on angiogenesis in vivo is limited. This study aimed to determine the role of DEPTOR in tissue angiogenesis and to elucidate the molecular mechanisms. METHODS: Cre/loxP conditional gene knockout strategy was used to delete the Deptor gene in mouse vascular ECs. The expression or distribution of cluster of differentiation 31 (CD31), vascular endothelial growth factor (VEGF) and hypoxia inducible factor-1 alpha (HIF-1α) were detected by immunohistochemical staining or western blot. Tube formation assay was used to measure angiogenesis in vitro. RESULTS: Deptor knockdown led to increased expression of CD31, VEGF and HIF-1α in heart, liver, kidney and aorta. After treatment with rapamycin, their expression was significantly down regulated. In vitro, human umbilical vein endothelial cells (HUVECs) were transfected with DEPTOR-specific small interfering RNA (siRNA), which resulted in a significant increase in endothelial tube formation and migration rates. In contrast, DEPTOR overexpression markedly reduced the expression of CD31, VEGF and HIF-1α. CONCLUSIONS: Our findings demonstrated that deletion of the Deptor gene in vascular ECs resulted in upregulated expression of CD31 and HIF-1α, and further stimulated the expression of VEGF which promoted angiogenesis, indicating that disruption of normal angiogenic pathways may occur through hyperactivation of the mTORC1/HIF-1α/VEGF signaling pathway.
BACKGROUND/AIMS: The mechanistic target of rapamycin (mTOR) signaling pathway is essential for angiogenesis and embryonic development. DEP domain-containing mTOR-interacting protein (DEPTOR) is an mTOR binding protein that functions to inhibit the mTOR pathway In vitro experiments suggest that DEPTOR is crucial for vascular endothelial cell (EC) activation and angiogenic responses. However, knowledge of the effects of DEPTOR on angiogenesis in vivo is limited. This study aimed to determine the role of DEPTOR in tissue angiogenesis and to elucidate the molecular mechanisms. METHODS: Cre/loxP conditional gene knockout strategy was used to delete the Deptor gene in mouse vascular ECs. The expression or distribution of cluster of differentiation 31 (CD31), vascular endothelial growth factor (VEGF) and hypoxia inducible factor-1 alpha (HIF-1α) were detected by immunohistochemical staining or western blot. Tube formation assay was used to measure angiogenesis in vitro. RESULTS:Deptor knockdown led to increased expression of CD31, VEGF and HIF-1α in heart, liver, kidney and aorta. After treatment with rapamycin, their expression was significantly down regulated. In vitro, human umbilical vein endothelial cells (HUVECs) were transfected with DEPTOR-specific small interfering RNA (siRNA), which resulted in a significant increase in endothelial tube formation and migration rates. In contrast, DEPTOR overexpression markedly reduced the expression of CD31, VEGF and HIF-1α. CONCLUSIONS: Our findings demonstrated that deletion of the Deptor gene in vascular ECs resulted in upregulated expression of CD31 and HIF-1α, and further stimulated the expression of VEGF which promoted angiogenesis, indicating that disruption of normal angiogenic pathways may occur through hyperactivation of the mTORC1/HIF-1α/VEGF signaling pathway.
Authors: Ahmad F Alghanem; Javier Abello; Joshua M Maurer; Ashutosh Kumar; Chau My Ta; Susheel K Gunasekar; Urooj Fatima; Chen Kang; Litao Xie; Oluwaseun Adeola; Megan Riker; Macaulay Elliot-Hudson; Rachel A Minerath; Chad E Grueter; Robert F Mullins; Amber N Stratman; Rajan Sah Journal: Elife Date: 2021-02-25 Impact factor: 8.140
Authors: Shan Wang; Ariel Raybuck; Eileen Shiuan; Sung Hoon Cho; Qingfei Wang; Dana M Brantley-Sieders; Deanna Edwards; Margaret M Allaman; James Nathan; Keith T Wilson; David DeNardo; Siyuan Zhang; Rebecca Cook; Mark Boothby; Jin Chen Journal: JCI Insight Date: 2020-08-06