Abstract
Background: Vascular activation is characterized by increased proinflammatory, pro thrombotic, and proadhesive signaling. Several chronic and acute conditions, including Bcr-abl-negative myeloproliferative neoplasms (MPNs), graft-vs-host disease, and COVID-19 have been noted to have increased activation of the janus kinase (JAK)-signal transducer and downstream activator of transcription (STAT) pathways. Two notable inhibitors of the JAK-STAT pathway are ruxolitinib (JAK1/2 inhibitor) and fedratinib (JAK2 inhibitor), which are currently used to treat MPN patients. However, in some conditions, it has been noted that JAK inhibitors can increase the risk of thromboembolic complications. Objectives: We sought to define the anti-inflammatory and antithrombotic effects of JAK-STAT inhibitors in vascular endothelial cells. Methods: We assessed endothelial activation in the presence or absence of ruxolitinib or fedratinib by using immunoblots, immunofluorescence, qRT-PCR, and function coagulation assays. Finally, we used endothelialized microfluidics perfused with blood from normal and JAK2V617F+ individuals to evaluate whether ruxolitinib and fedratinib changed cell adhesion. Results: We found that both ruxolitinib and fedratinib reduced endothelial cell phospho-STAT1 and STAT3 signaling and attenuated nuclear phospho-NK-κB and phospho-c-Jun localization. JAK-STAT inhibition also limited secretion of proadhesive and procoagulant P-selectin and von Willebrand factor and proinflammatory IL-6. Likewise, we found that JAK-STAT inhibition reduced endothelial tissue factor and urokinase plasminogen activator expression and activity. Conclusions: By using endothelialized microfluidics perfused with whole blood samples, we demonstrated that endothelial treatment with JAK-STAT inhibitors prevented rolling of both healthy control and JAK2V617F MPN leukocytes. Together, these findings demonstrate that JAK-STAT inhibitors reduce the upregulation of critical prothrombotic pathways and prevent increased leukocyte–endothelial adhesion.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 1366-1380 |
| Number of pages | 15 |
| Journal | Journal of Thrombosis and Haemostasis |
| Volume | 21 |
| Issue number | 5 |
| DOIs | |
| State | Published - May 2023 |
Bibliographical note
Funding Information:J.D.B. receives funds from Bayer independent from work herein. G.M.V. receives research funding from CSL Behring and Mitobridge (Astellas). C.B. has received reagents from CTI BioPharma for the conduct of clinical trial NCT 02891603 and a pending patent, WO2017058950A1, for methods of treating transplant rejection. B.C.B. holds patents related to CD4+ T cell pSTAT3 as a marker and therapeutic target of acute GVHD (WO2015120436A2); for the use of JAK inhibitors for rejection and GVHD prevention (WO2017058950A1); and for the use of CD83-targeted chimeric antigen receptor T cells in GVHD prevention, immune tolerance, autoimmunity, and acute myeloid leukemia therapy (WO2019165156). At this time, neither B.C.B. nor the University of Minnesota has received payment related to claims described in the patent. B.C.B. has received honoraria for participating in advisory board discussions for Incyte Corp and CTI BioPharma within the past 5 years. Remaining authors have no conflicts to disclose.
Funding Information:
Funding information J.D.B. is supported in part by NHLBI K08HL159289, Institutional Research Grant #129819-IRG-16-189-58-IRG-114 from the American Cancer Society , Masonic Cancer Research Translational Initiative Research Grant, an American Heart Association Career Development Award and American Society of Hematology Restart Award. A.D.S. is supported by T32-HL139341. D.R. received funding from University of Minnesota Life Sciences Undergraduate Research Program (LSSURP) 5R25HL088728-14. Academic Investment in Research Program Grant supports G.M.V. D.K.W. is supported by R01HL140589 and HL132906. Portions of this work were conducted in the Minnesota Nano Center, which is supported by the National Science Foundation through the National Nanotechnology Coordinated Infrastructure (NNCI) under Award Number ECCS-2025124. Other portions were supported by the resources and staff at the University of Minnesota University Imaging Centers (UIC), SCR_020997. Database and Biostatistic support was provided by Clinical and Translational Science Institute grant support (UL1TR002494 from the National Institutes of Health’s National Center for Advancing Translational Sciences).
Funding Information:
J.D.B. is supported in part by NHLBI K08HL159289, Institutional Research Grant #129819-IRG-16-189-58-IRG-114 from the American Cancer Society, Masonic Cancer Research Translational Initiative Research Grant, an American Heart Association Career Development Award and American Society of Hematology Restart Award. A.D.S. is supported by T32-HL139341. D.R. received funding from University of Minnesota Life Sciences Undergraduate Research Program (LSSURP) 5R25HL088728-14. Academic Investment in Research Program Grant supports G.M.V. D.K.W. is supported by R01HL140589 and HL132906. Portions of this work were conducted in the Minnesota Nano Center, which is supported by the National Science Foundation through the National Nanotechnology Coordinated Infrastructure (NNCI) under Award Number ECCS-2025124. Other portions were supported by the resources and staff at the University of Minnesota University Imaging Centers (UIC), SCR_020997. Database and Biostatistic support was provided by Clinical and Translational Science Institute grant support (UL1TR002494 from the National Institutes of Health’s National Center for Advancing Translational Sciences).
Funding Information:
Funding information J.D.B. is supported in part by NHLBI K08HL159289, Institutional Research Grant #129819-IRG-16-189-58-IRG-114 from the American Cancer Society, Masonic Cancer Research Translational Initiative Research Grant, an American Heart Association Career Development Award and American Society of Hematology Restart Award. A.D.S. is supported by T32-HL139341. D.R. received funding from University of Minnesota Life Sciences Undergraduate Research Program (LSSURP) 5R25HL088728-14. Academic Investment in Research Program Grant supports G.M.V. D.K.W. is supported by R01HL140589 and HL132906. Portions of this work were conducted in the Minnesota Nano Center, which is supported by the National Science Foundation through the National Nanotechnology Coordinated Infrastructure (NNCI) under Award Number ECCS-2025124. Other portions were supported by the resources and staff at the University of Minnesota University Imaging Centers (UIC), SCR_020997. Database and Biostatistic support was provided by Clinical and Translational Science Institute grant support (UL1TR002494 from the National Institutes of Health's National Center for Advancing Translational Sciences).The authors would like to thank Michael Franklin, MS for editorial assistance in manuscript preparation. The authors would like to thank Muse Jama, Amy Eisenberg, Diondra Howard, Ghislaine Feussom, and Rebecca Cote for assistance with acquiring whole blood samples from volunteers. The authors also thank Drs. Nigel Key, John Belcher, and Robert Hebbel for helpful comments and insights. J.D.B. designed and performed experiments, analyzed data, and wrote the manuscript. A.D.S. designed and performed experiments, analyzed data, and edited the manuscript. E.A. performed experiments, analyzed data, and edited the manuscript. A.N. and J.N. performed experiments and analyzed data. G.H. and D.R. designed and performed experiments and analyzed data. G.M.V. B.C.B. and D.K.W. designed experiments and edited the manuscript. J.D.B. receives funds from Bayer independent from work herein. G.M.V. receives research funding from CSL Behring and Mitobridge (Astellas). C.B. has received reagents from CTI BioPharma for the conduct of clinical trial NCT 02891603 and a pending patent, WO2017058950A1, for methods of treating transplant rejection. B.C.B. holds patents related to CD4+ T cell pSTAT3 as a marker and therapeutic target of acute GVHD (WO2015120436A2); for the use of JAK inhibitors for rejection and GVHD prevention (WO2017058950A1); and for the use of CD83-targeted chimeric antigen receptor T cells in GVHD prevention, immune tolerance, autoimmunity, and acute myeloid leukemia therapy (WO2019165156). At this time, neither B.C.B. nor the University of Minnesota has received payment related to claims described in the patent. B.C.B. has received honoraria for participating in advisory board discussions for Incyte Corp and CTI BioPharma within the past 5 years. Remaining authors have no conflicts to disclose. J.D.B. is supported in part by NHLBI K08HL159289, Institutional Research Grant #129819-IRG-16-189-58-IRG-114 from the American Cancer Society, Masonic Cancer Research Translational Initiative Research Grant, an American Heart Association Career Development Award and American Society of Hematology Restart Award. A.D.S. is supported by T32-HL139341. D.R. received funding from University of Minnesota Life Sciences Undergraduate Research Program (LSSURP) 5R25HL088728-14. Academic Investment in Research Program Grant supports G.M.V. D.K.W. is supported by R01HL140589 and HL132906. Portions of this work were conducted in the Minnesota Nano Center, which is supported by the National Science Foundation through the National Nanotechnology Coordinated Infrastructure (NNCI) under Award Number ECCS-2025124. Other portions were supported by the resources and staff at the University of Minnesota University Imaging Centers (UIC), SCR_020997. Database and Biostatistic support was provided by Clinical and Translational Science Institute grant support (UL1TR002494 from the National Institutes of Health's National Center for Advancing Translational Sciences).
Publisher Copyright:
© 2023 The Author(s)
Keywords
- endothelium
- janus kinase inhibitors
- myeloproliferative neoplasm
- thrombosis
- tissue factor
