Elucidating PAR1 as a therapeutic target for delayed traumatic brain injury: Unveiling the PPAR-γ/Nrf2/HO-1/GPX4 axis to suppress ferroptosis and alleviate NLRP3 inflammasome activation in rats

Article

Author: Abdallah, Dalaal M ; Soubh, Ayman A ; El-Gazar, Amira A ; El-Abhar, Hanan S ; Ragab, Ghada M

Repetitive traumatic brain injury (RTBI) is acknowledged as a silent overlooked public health crisis, with an incomplete understanding of its pathomechanistic signaling pathways. Mounting evidence suggests the involvement of thrombin and its receptor, the protease-activated receptor (PAR)1, in the development of secondary injury in TBI; however, the consequences of PAR1 modulation and its impact on ferroptosis-redox signaling, and NLRP3 inflammasome activation in RTBI, remain unclear. Further, the utilitarian function of PAR1 as a therapeutic target in RTBI has not been elucidated. To study this crosstalk, RTBI was induced in Wistar rats by daily weight drops on the right frontal region for five days. Three groups were included: normal control, untreated RTBI, and RTBI+SCH79797 (a PAR1 inhibitor administered post-trauma at 25 μg/kg/day). The concomitant treatment of PAR1 antagonism improved altered behavior function, cortical histoarchitecture, and neuronal cell survival. Moreover, the receptor blockade downregulated mRNA expression of PAR1 but upregulatedthat of the neuroprotective receptor PPAR-γ. The anti-inflammatory impact of SCH79797 was signified by the low immune expression/levels of NF-κB p65,TNF-α, IL-1β, and IL-18. Consequently, the PAR1 blocker hindered the formation of inflammasome components NLRP3, ASC, and activated caspase-1. Ultimately, SCH79797 treatment abated ferroptosis-dependent iron redox signaling through the activation of the antioxidant Nrf2/HO-1 axis and its subsequent antioxidant machinery (GPX4, SOD) to limit lipid peroxidation, iron accumulation, and transferrin serum increment. Collectively, SCH79797 offered putative preventive mechanisms against secondary RTBI consequences in rats by impeding ferroptosis and NLRP3 inflammasome through activating the PPAR-γ/Nrf2 antioxidant cue.

14 Nov 2019·Mediators of InflammationQ3 · MEDICINE

Thrombin Induces Secretion of Multiple Cytokines and Expression of Protease-Activated Receptors in Mouse Mast Cell Line

Q3 · MEDICINE

ArticleOA

Author: Yu, Yingyan ; Zhu, Tao ; Guo, Zaipei ; Liao, Ren ; Li, Jingyi ; Fang, Xiaobin

Background. Thrombin could elicit degranulation of mast cells involved in numerous physiologic and pathologic processes; however, the detailed scrutiny of this procedure and further research of possible cell signaling pathways are lacking.Methods. P815 mouse mast cells were exposed to various concentrations of thrombin for 16 h. Expression of protease-activated receptor (PAR)1, PAR2, PAR3, and PAR4 mRNA in P815 was analyzed by quantitative real-time PCR (qRT-PCR) and the fittest concentration of thrombin was decided. Then, secretions of mediators from P815 stimulated by thrombin 0.2 U/ml were determined using enzyme-linked immunosorbent assay (ELISA) and Luminex liquichip; the possible cell signaling pathways were measured by immunoblotting. Furthermore, inhibition of thrombin inhibitor (hirudin), PAR1 inhibitor (SCH79797), and MAPK inhibitors (SB203580, PD98059, and SP600125) on the mediator section was evaluated by ELISA and Luminex liquichip.Results. Thrombin 0.2 U/ml induced the elevated expression of PAR1, PAR2, PAR3, and PAR4, as well as the increasing level of phospho-IκBα, phospho-SAPK/JNK MAPK, phospho-P38 MAPK (Thr180/Tyr182), and phospho-ERK1/2 MAPK (p44/42) in P815. Secretion of vascular endothelial growth factor (VEGF), tumor necrosis factor-α(TNF-α), interleukin- (IL-) 2, IL-6, chemokine ligand- (CCL-) 2, chemokine (C-X-C motif) ligand- (CXCL-) 1, and CXCL-5 from P815 increased apparently; this effect could be diminished by hirudin, whereas SCH79797 and MAPK inhibitors (SB203580, PD98059, and SP600125) diminish the secretions with weaker effect.Conclusion. We found the expression of PAR mRNA in P815, activation of signaling pathways of nuclear factor-kappaB (NF-κB), and mitogen-activated protein kinases (MAPKs) including C-Jun NH2-terminal kinase (JNK), P38, and extracellular signal-regulated kinase 1/2 (ERK1/2), and the release of multiple inflammatory mediators stimulated by thrombin, as well as the inhibition of the inflammatory releases by hirudin, SCH79797, and MAPK inhibitors including SB203580, PD98059, and SP600125.

01 Mar 2016·Experimental Biology and MedicineQ4 · MEDICINE

Featured Article: Differential regulation of endothelial nitric oxide synthase phosphorylation by protease-activated receptors in adult human endothelial cells

Q4 · MEDICINE

Article

Author: Motley, Evangeline D ; Eguchi, Satoru ; Tillery, Lakeisha C ; Epperson, Tenille A

Protease-activated receptors have been shown to regulate endothelial nitric oxide synthase through the phosphorylation of specific sites on the enzyme. It has been established that PAR-2 activation phosphorylates eNOS-Ser-1177 and leads to the production of the potent vasodilator nitric oxide, while PAR-1 activation phosphorylates eNOS-Thr-495 and decreases nitric oxide production in human umbilical vein endothelial cells. In this study, we hypothesize a differential coupling of protease-activated receptors to the signaling pathways that regulates endothelial nitric oxide synthase and nitric oxide production in primary adult human coronary artery endothelial cells. Using Western Blot analysis, we showed that thrombin and the PAR-1 activating peptide, TFLLR, lead to the phosphorylation of eNOS-Ser-1177 in human coronary artery endothelial cells, which was blocked by SCH-79797 (SCH), a PAR-1 inhibitor. Using the nitrate/nitrite assay, we also demonstrated that the thrombin- and TFLLR-induced production of nitric oxide was inhibited by SCH and L-NAME, a NOS inhibitor. In addition, we observed that TFLLR, unlike thrombin, significantly phosphorylated eNOS-Thr-495, which may explain the observed delay in nitric oxide production in comparison to that of thrombin. Activation of PAR-2 by SLIGRL, a PAR-2 specific ligand, leads to dual phosphorylation of both catalytic sites but primarily regulated eNOS-Thr-495 phosphorylation with no change in nitric oxide production in human coronary artery endothelial cells. PAR-3, known as the non-signaling receptor, was activated by TFRGAP, a PAR-3 mimicking peptide, and significantly induced the phosphorylation of eNOS-Thr-495 with minimal phosphorylation of eNOS-Ser-1177 with no change in nitric oxide production. In addition, we confirmed that PAR-mediated eNOS-Ser-1177 phosphorylation was Ca2+-dependent using the Ca2+ chelator, BAPTA, while eNOS-Thr-495 phosphorylation was mediated via Rho kinase using the ROCK inhibitor, Y-27632, suggesting protease-activated receptor coupling to Gq and G12/13, respectively. These data suggest a vascular bed specific differential coupling of protease-activated receptors to the signaling pathways that regulate endothelial nitric oxide synthase and nitric oxide production that may be responsible for endothelial dysfunction associated with cardiovascular disease.

100 Deals associated with SCH-79797

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Indication	Highest Phase	Country/Location	Organization	Date
Gram-Negative Bacterial Infections	Phase 2	US	Princeton University	25 Jun 2020
Gram-Positive Bacterial Infections	Phase 2	US	Princeton University	25 Jun 2020
Oxygen radical damage	Preclinical	AU	The University of New South Wales	03 Nov 2015

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