Baorong Li1, Yingmiao Liu1, Tenglong Hu2,3, Yan Zhang4, Cong Zhang5, Tao Li4, Chunxu Wang4, Zengxiang Dong6, Valerie A Novakovic7, Tianshui Hu4, Jialan Shi8,9. 1. Department of Stomatology, The First Hospital, Harbin Medical University, Harbin, 150001, China. 2. Department of Stomatology, The First Hospital, Harbin Medical University, Harbin, 150001, China. hutenglong2018net@163.com. 3. Department of Oral Anatomy and Physiology, Stomatology School, Harbin Medical University, Harbin, 150001, China. hutenglong2018net@163.com. 4. Department of Hematology, The First Hospital, Harbin Medical University, Harbin, 150001, China. 5. Department of Ultrasound, The First Hospital, Harbin Medical University, Harbin, 150001, China. 6. Department of Cardiology, The First Hospital, Harbin Medical University, Harbin, 150001, China. 7. Department of Research and Surgery, VA Boston Healthcare System, Brigham and Women's Hospital, Harvard Medical School, Boston, 02132, USA. 8. Department of Hematology, The First Hospital, Harbin Medical University, Harbin, 150001, China. jialan_shi@hms.harvard.edu. 9. Department of Research and Surgery, VA Boston Healthcare System, Brigham and Women's Hospital, Harvard Medical School, Boston, 02132, USA. jialan_shi@hms.harvard.edu.
Abstract
BACKGROUND: Hypercoagulability is a major cancer-associated complication linked to poor patient prognosis. The production of neutrophil extracellular traps (NETs) is increasingly found to be linked with the development and metastasis of cancer, as well as with thrombi formation in cancer patients. We hypothesized that the neutrophil NET release may be triggered by specific cytokines in oral squamous cell carcinoma (OSCC) patients, thereby predisposing them to a hypercoagulable state. Moreover, we have evaluated the interaction between NETs and endothelial cells (ECs). METHODS: NET procoagulant activity was assessed based on fibrin and purified coagulation complex production assays, as well as by measuring coagulation time (CT). We further used confocal microscopy to quantify the exposure of phosphatidylserine (PS), fibrin strands, and cell FVa/Xa binding. RESULTS: OSCC patients with stage III/IV exhibited elevated plasma NET levels compared to stage I/II or CTR (all P < 0.05). Neutrophils from OSCC patients are predisposed to amplified NET release compared to those from CTR. Furthermore, depleting IL-8, IL-6, and TNF-α led to a reduction in NET release in the plasma. OSCC NETs increased thrombin and fibrin generation and decreased CT significantly (P < 0.05). When NETs were isolated and used to treat ECs, these cells exhibited disrupted morphology by retracting from their cell-cell junctions and convert to a procoagulant phenotype. These effects could be attenuated by approximately 70% using DNase I. CONCLUSIONS: Our findings are consistent with a model wherein OSCC drives a systemic inflammatory state, which, in turn, drives neutrophils to prime and release NETs, which drive the development of a hypercoagulable state. Intervening in this process may be a viable means of disrupting these undesirable coagulation dynamics in stage III/IV OSCC patients.
BACKGROUND: Hypercoagulability is a major cancer-associated complication linked to poor patient prognosis. The production of neutrophil extracellular traps (NETs) is increasingly found to be linked with the development and metastasis of cancer, as well as with thrombi formation in cancerpatients. We hypothesized that the neutrophil NET release may be triggered by specific cytokines in oral squamous cell carcinoma (OSCC) patients, thereby predisposing them to a hypercoagulable state. Moreover, we have evaluated the interaction between NETs and endothelial cells (ECs). METHODS: NET procoagulant activity was assessed based on fibrin and purified coagulation complex production assays, as well as by measuring coagulation time (CT). We further used confocal microscopy to quantify the exposure of phosphatidylserine (PS), fibrin strands, and cell FVa/Xa binding. RESULTS: OSCC patients with stage III/IV exhibited elevated plasma NET levels compared to stage I/II or CTR (all P < 0.05). Neutrophils from OSCC patients are predisposed to amplified NET release compared to those from CTR. Furthermore, depleting IL-8, IL-6, and TNF-α led to a reduction in NET release in the plasma. OSCC NETs increased thrombin and fibrin generation and decreased CT significantly (P < 0.05). When NETs were isolated and used to treat ECs, these cells exhibited disrupted morphology by retracting from their cell-cell junctions and convert to a procoagulant phenotype. These effects could be attenuated by approximately 70% using DNase I. CONCLUSIONS: Our findings are consistent with a model wherein OSCC drives a systemic inflammatory state, which, in turn, drives neutrophils to prime and release NETs, which drive the development of a hypercoagulable state. Intervening in this process may be a viable means of disrupting these undesirable coagulation dynamics in stage III/IV OSCC patients.