PMID- 31831282 OWN - NLM STAT- MEDLINE DCOM- 20201211 LR - 20201214 IS - 1873-569X (Electronic) IS - 0923-1811 (Linking) VI - 97 IP - 1 DP - 2020 Jan TI - Rip 1-dependent endothelial necroptosis participates in ischemia-reperfusion injury of mouse flap. PG - 30-40 LID - S0923-1811(19)30364-0 [pii] LID - 10.1016/j.jdermsci.2019.11.009 [doi] AB - BACKGROUND: Ischemia reperfusion injury plays an important role in free flap necrosis. However, the detailed mechanism is not clear, and effective methods for improving the survival rate of skin flap are still lacking. OBJECTIVE: To investigate the regulation and functional link between necroptosis and ischemia-reperfusion injury of mouse flap. METHODS: We established a mouse ischemia-reperfusion injury flap model and a cell Oxygen Glucose Deprivation (OGD) model intervened with Necrostatin-1. The mouse flap tissues were harvested in vivo for histological immunofluorescence analysis and western blotting analyses. The HUVECs cells with various treatments in vitro were assessed by using Transwell assay, tube formation assay, cell counting kit-8 analysis and flow cytometry. A Rip3-knockout cell line and a TNFR1-knockout cell line were generated from HUVEC cells using the CRISPR-Cas9 technology and were subsequently used to explore the related mechanisms. RESULTS: The expression of p-Rip3 is positive in both mouse and cell culture models. When necroptosis is completely or partially inhibited in vivo, damaged tissues are repaired with better efficiency. The cells treated with Necrostatin-1 in vitro exhibit faster migration, proliferation and better tube formation. Deficiency of TNFR1 can block the necroptosis pathway by blocking the phosphorylation of Rip3 in HUVEC OGD/ROG model. Meanwhile, the levels of APJ, HIF-1alpha, and VEGF are reduced when necroptosis is inhibited by Necrostatin-1. CONCLUSION: TNFR1 mediates Rip1/Rip3 in ischemia-reperfusion injury. Inhibition of necroptosis attenuates the ischemia-reperfusion injury of flap and may enhance hypoxic tolerance of HUVECs and vascular homeostasis through regulation of the HIF-1alpha signaling pathways. CI - Copyright (c) 2019 Japanese Society for Investigative Dermatology. Published by Elsevier B.V. All rights reserved. FAU - Geng, Lele AU - Geng L AD - Department of Plastic Surgery, Shanghai Ninth People's Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, China. FAU - Zhang, Gai AU - Zhang G AD - Department of Plastic Surgery, Shanghai Ninth People's Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, China. FAU - Yao, Min AU - Yao M AD - Department of Plastic Surgery, Shanghai Ninth People's Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, China. FAU - Fang, Yong AU - Fang Y AD - Department of Plastic Surgery, Shanghai Ninth People's Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, China. Electronic address: fangyong1020@hotmail.com. LA - eng PT - Journal Article DEP - 20191128 PL - Netherlands TA - J Dermatol Sci JT - Journal of dermatological science JID - 9011485 RN - 0 (Imidazoles) RN - 0 (Indoles) RN - 0 (Receptors, Tumor Necrosis Factor, Type I) RN - 0 (TNFRSF1A protein, human) RN - 0 (Tnfrsf1a protein, mouse) RN - 0 (necrostatin-1) RN - EC 2.7.11.1 (RIPK3 protein, human) RN - EC 2.7.11.1 (Receptor-Interacting Protein Serine-Threonine Kinases) RN - EC 2.7.11.1 (Ripk1 protein, mouse) SB - IM MH - Animals MH - Cell Hypoxia/drug effects MH - Disease Models, Animal MH - Endothelium, Vascular/drug effects/pathology MH - Free Tissue Flaps/blood supply/*pathology/transplantation MH - Gene Knockdown Techniques MH - Graft Survival/*drug effects MH - HEK293 Cells MH - Human Umbilical Vein Endothelial Cells MH - Humans MH - Imidazoles/*administration & dosage MH - Indoles/*administration & dosage MH - Injections, Intravenous MH - Male MH - Mice MH - Necroptosis/*drug effects/physiology MH - Phosphorylation MH - Receptor-Interacting Protein Serine-Threonine Kinases/genetics/metabolism MH - Receptors, Tumor Necrosis Factor, Type I/genetics/metabolism MH - Reperfusion Injury/pathology/*prevention & control MH - Signal Transduction/drug effects/genetics OTO - NOTNLM OT - Ischemia reperfusion injury OT - Necroptosis OT - Necrostatin-1 COIS- Declaration of Competing Interest The authors have declared no conflict of interest. EDAT- 2019/12/14 06:00 MHDA- 2020/12/15 06:00 CRDT- 2019/12/14 06:00 PHST- 2019/07/19 00:00 [received] PHST- 2019/11/21 00:00 [revised] PHST- 2019/11/24 00:00 [accepted] PHST- 2019/12/14 06:00 [pubmed] PHST- 2020/12/15 06:00 [medline] PHST- 2019/12/14 06:00 [entrez] AID - S0923-1811(19)30364-0 [pii] AID - 10.1016/j.jdermsci.2019.11.009 [doi] PST - ppublish SO - J Dermatol Sci. 2020 Jan;97(1):30-40. doi: 10.1016/j.jdermsci.2019.11.009. Epub 2019 Nov 28.