Activation of NLRP3 inflammasome complex potentiates venous thrombosis in response to hypoxia.
Gupta Neha,Sahu Anita,Prabhakar Amit,Chatterjee Tathagata,Tyagi Tarun,Kumari Babita,Khan Nilofar,Nair Velu,Bajaj Nitin,Sharma Manish,Ashraf Mohammad Zahid
Proceedings of the National Academy of Sciences of the United States of America
Venous thromboembolism (VTE), caused by altered hemostasis, remains the third most common cause of mortality among all cardiovascular conditions. In addition to established genetic and acquired risk factors, low-oxygen environments also predispose otherwise healthy individuals to VTE. Although disease etiology appears to entail perturbation of hemostasis pathways, the key molecular determinants during immediate early response remain elusive. Using an established model of venous thrombosis, we here show that systemic hypoxia accelerates thromboembolic events, functionally stimulated by the activation of nucleotide binding domain, leucine-rich-containing family, pyrin domain containing 3 (NLRP3) inflammasome complex and increased IL-1β secretion. Interestingly, we also show that the expression of NLRP3 is mediated by hypoxia-inducible factor 1-alpha (HIF-1α) during these conditions. The pharmacological inhibition of caspase-1, in vivo knockdown of NLRP3, or HIF-1α other than IL-1β-neutralizing antibodies attenuated inflammasome activation and curtailed thrombosis under hypoxic conditions. We extend the significance of these preclinical findings by studying modulation of this pathway in patients with altitude-induced venous thrombosis. Our results demonstrate distinctive, increased expression of NLRP3, caspase-1, and IL-1β in individuals with clinically established venous thrombosis. We therefore propose that an early proinflammatory state in the venous milieu, orchestrated by the HIF-induced NLRP3 inflammasome complex, is a key determinant of acute thrombotic events during hypoxic conditions.
10.1073/pnas.1620458114
Endoplasmic reticulum stress and oxidative stress contribute to neuronal pyroptosis caused by cerebral venous sinus thrombosis in rats: Involvement of TXNIP/peroxynitrite-NLRP3 inflammasome activation.
Ding Rui,Ou Weiyang,Chen Chengwei,Liu Yaqi,Li Haiyan,Zhang Xifang,Chai Huihui,Ding Xiaowen,Wang Qiujing
Neurochemistry international
Cerebral venous sinus thrombosis (CVST) is a rare type of stroke, which is life-threatening in severe cases. However, considerably less attention has been concentrated on the mechanism of neural cell damage after CVST. This study aims to investigate the role of endoplasmic reticulum stress, oxidative stress, and pyroptosis in a well-established rodent model of CVST. Rat brains were harvested at 0 h, 6 h, days 1, days 3, days 7, and days 14 post-CVST for measurement of corresponding indexes. Endoplasmic reticulum stress sensors (including protein kinase RNA-like ER kinase (PERK) and inositol-requiring enzyme-1α (IRE1α)), oxidative stress markers (thioredoxin-interacting protein (TXNIP) and peroxynitrite), NLRP3, caspase p20, IL-1β, and gasdermin D (GSDMD, an indicator of pyroptosis) were separately evaluated by Western-blot and Immunohistochemistry/Immunofluorescence. Co-immunoprecipitation and Fluorescent double-labeling were employed to probe into the relationship between TXNIP/peroxynitrite and NLRP3 inflammasome. In the damaged cortex region, profuse p-PERK, p-IRE1α, TXNIP were produced and predominantly localized in neurons accompanied by a small amount expressed in microglia and astrocytes. The levels of 3-nitrotyrosine (3-NT, as a footprint of peroxynitrite), NLRP3, caspase p20, IL-1β, and GSDMD were distinctly elevated post-CVST and cellular localization of peroxynitrite, NLRP3, caspase p20, and IL-1β was largely observed in neurons and/or microglia. Importantly, sites of enhanced TXNIP and 3-NT immunoreactivity were colocalized with increased NLRP3 staining, indicating the involvement of TXNIP and peroxynitrite in NLRP3 inflammasome activation and subsequent pyroptosis. Besides, co-immunoprecipitation also hinted that there might be an interaction or causality between TXNIP/peroxynitrite and NLRP3 inflammasome. We concluded that endoplasmic reticulum stress and oxidative stress may jointly lead to neuronal NLRP3 inflammasome activation and pyroptosis after CVST.
10.1016/j.neuint.2020.104856
Inflammatory Mediators of Endothelial Dysfunction.
Life (Basel, Switzerland)
Endothelial dysfunction (ED) is characterized by imbalanced vasodilation and vasoconstriction, elevated reactive oxygen species (ROS), and inflammatory factors, as well as deficiency of nitric oxide (NO) bioavailability. It has been reported that the maintenance of endothelial cell integrity serves a significant role in human health and disease due to the involvement of the endothelium in several processes, such as regulation of vascular tone, regulation of hemostasis and thrombosis, cell adhesion, smooth muscle cell proliferation, and vascular inflammation. Inflammatory modulators/biomarkers, such as IL-1α, IL-1β, IL-6, IL-12, IL-15, IL-18, and tumor necrosis factor α, or alternative anti-inflammatory cytokine IL-10, and adhesion molecules (ICAM-1, VCAM-1), involved in atherosclerosis progression have been shown to predict cardiovascular diseases. Furthermore, several signaling pathways, such as NLRP3 inflammasome, that are associated with the inflammatory response and the disrupted H2S bioavailability are postulated to be new indicators for endothelial cell inflammation and its associated endothelial dysfunction. In this review, we summarize the knowledge of a plethora of reviews, research articles, and clinical trials concerning the key inflammatory modulators and signaling pathways in atherosclerosis due to endothelial dysfunction.
10.3390/life13061420
Gegen Qinlian pills alleviate carrageenan-induced thrombosis in mice model by regulating the HMGB1/NF-κB/NLRP3 signaling.
Phytomedicine : international journal of phytotherapy and phytopharmacology
BACKGROUND:The high incidence of thrombotic events is one of the clinical characteristics of coronavirus disease of 2019 (COVID-19), due to a hyperinflammatory response caused by the virus. Gegen Qinlian Pills (GQP) is a Traditional Chinese Medicine that is included in the Chinese Pharmacopoeia and played an important role in the clinical fight against COVID-19. Although GQP has shown the potential to treat thrombosis, there is no relevant research on its treatment of thrombosis so far. HYPOTHESIS:We hypothesized that GQP may be capable inhibit inflammation-induced thrombosis. STUDY DESIGN:We tested our hypothesis in a carrageenan-induced thrombosis mouse model in vivo and lipopolysaccharide (LPS)-induced human endothelial cells (HUVECs) in vitro. METHODS:We used a carrageenan-induced mouse thrombus model to confirm the inhibitory effect of GQP on inflammation-induced thrombus. In vitro, studies in human umbilical vein endothelial cells (HUVECs) and in silico network pharmacology analyses were performed to reveal the underlying mechanisms of GQP and determine the main components, targets, and pathways of GQP, respectively. RESULTS:Oral administration of 227.5 mg/kg, 445 mg/kg and 910 mg/kg of GQP significantly inhibited thrombi in the lung, liver, and tail and augmented tail blood flow of carrageenan-induced mice with reduced plasma tumor necrosis factor α (TNF-α) and diminished expression of high mobility group box 1 (HMGB1) in lung tissues. GQP ethanol extract (1, 2, or 5 μg/ml) also reduced the adhesion of platelets to LPS stimulated HUVECs. The TNF-α and the expression of HMGB1, nuclear factor kappa B (NF-κB), and NLR family pyrin domain containing 3 (NLRP3) in LPS stimulated HUVECs were also attenuated. Moreover, we analyzed the components of GQP and inferred the main targets, biological processes, and pathways of GQP in the treatment of inflammation-induced thrombosis through network pharmacology. CONCLUSION:Overall, we demonstrated that GQP could reduce inflammation-induced thrombosis by inhibiting HMGB1/NFκB/NLRP3 signaling and provided an accurate explanation for the multi-target, multi-function mechanism of GQP in the treatment of thromboinflammation, and provides a reference for the clinical usage of GQP.
10.1016/j.phymed.2022.154083
TXNIP activates NLRP3/IL-1β and participate in inflammatory response and oxidative stress to promote deep venous thrombosis.
Experimental biology and medicine (Maywood, N.J.)
Increasing evidence indicates that deep venous thrombosis (DVT) is a common peripheral vascular disease. This study aims to investigate the mechanisms of thioredoxin-interacting protein (TXNIP) and nod-like receptor protein 3 (NLRP3) inflammasome in deep venous thrombosis (DVT). A total of 66 Sprague-Dawley (SD) rats were employed to conduct DVT model. DVT rat was treated with silenced TXNIP (si-TXNIP) lentivirus and MCC950 (a NLRP3 inhibitor). The thrombosis weight and weight/length ratio, tissue factor, inflammatory factors, superoxide dismutase (SOD), malondialdehyde (MDA), and glutathione peroxidase (GSH-Px) were measured. Hematoxylin-eosin (H&E) staining was used to investigate the pathological change. Western blotting was used to determine the protein expression level. The expression level of thioredoxin (TRx) was suppressed, whereas TXNIP and NLRP3 were elevated in DVT rat. Si-TXNIP or MCC950 could reduce the thrombosis weight and weight/length ratio, ameliorate the pathological change, and decrease inflammatory reaction. Mechanistically, si-TXNIP or MCC950 inhibited the expression levels of TXNIP, NLRP3, and interleukin (IL)-1β while elevating the TRx level, thereby suppressing the DVT. Our study indicated that si-TXNIP or MCC950 injection rescued the injury of vein induced by DVT. The possible mechanisms connected with the inhibition of TXNIP and NLRP3. TXNIP is a possible therapeutic target for DVT.
10.1177/15353702231191124
Inflammasome Signaling, Thromboinflammation, and Venous Thromboembolism.
JACC. Basic to translational science
Venous thromboembolism (VTE) remains a major health burden despite anticoagulation advances, suggesting incomplete management of pathogenic mechanisms. The NLRP3 (NACHT-, LRR- and pyrin domain-containing protein 3) inflammasome, interleukin (IL)-1, and pyroptosis are emerging contributors to the inflammatory pathogenesis of VTE. Inflammasome pathway activation occurs in patients with VTE. In preclinical models, inflammasome signaling blockade reduces venous thrombogenesis and vascular injury, suggesting that this therapeutic approach may potentially maximize anticoagulation benefits, protecting from VTE occurrence, recurrence, and ensuing post-thrombotic syndrome. The nonselective NLRP3 inhibitor colchicine and the anti-IL-1β agent canakinumab reduce atherothrombosis without increasing bleeding. Rosuvastatin reduces primary venous thrombotic events at least in part through lipid-lowering independent mechanisms, paving the way to targeted anti-inflammatory strategies in VTE. This review outlines recent preclinical and clinical evidence supporting a role for inflammasome pathway activation in venous thrombosis, and discusses the, yet unexplored, therapeutic potential of modulating inflammasome signaling to prevent and manage VTE.
10.1016/j.jacbts.2023.03.017
Inflammatory Mechanisms Contributing to Endothelial Dysfunction.
Theofilis Panagiotis,Sagris Marios,Oikonomou Evangelos,Antonopoulos Alexios S,Siasos Gerasimos,Tsioufis Costas,Tousoulis Dimitris
Biomedicines
Maintenance of endothelial cell integrity is an important component of human health and disease since the endothelium can perform various functions including regulation of vascular tone, control of hemostasis and thrombosis, cellular adhesion, smooth muscle cell proliferation, and vascular inflammation. Endothelial dysfunction is encompassed by complex pathophysiology that is based on endothelial nitric oxide synthase uncoupling and endothelial activation following stimulation from various inflammatory mediators (molecular patterns, oxidized lipoproteins, cytokines). The downstream signaling via nuclear factor-κB leads to overexpression of adhesion molecules, selectins, and chemokines that facilitate leukocyte adhesion, rolling, and transmigration to the subendothelial space. Moreover, oscillatory shear stress leads to pro-inflammatory endothelial activation with increased monocyte adhesion and endothelial cell apoptosis, an effect that is dependent on multiple pathways and flow-sensitive microRNA regulation. Moreover, the role of neutrophil extracellular traps and NLRP3 inflammasome as inflammatory mechanisms contributing to endothelial dysfunction has recently been unveiled and is under further investigation. Consequently, and following their activation, injured endothelial cells release inflammatory mediators and enter a pro-thrombotic state through activation of coagulation pathways, downregulation of thrombomodulin, and an increase in platelet adhesion and aggregation owing to the action of von-Willebrand factor, ultimately promoting atherosclerosis progression.
10.3390/biomedicines9070781
Glucose metabolite methylglyoxal induces vascular endothelial cell pyroptosis via NLRP3 inflammasome activation and oxidative stress in vitro and in vivo.
Cellular and molecular life sciences : CMLS
Methylglyoxal (MGO), a reactive dicarbonyl metabolite of glucose, plays a prominent role in the pathogenesis of diabetes and vascular complications. Our previous studies have shown that MGO is associated with increased oxidative stress, inflammatory responses and apoptotic cell death in endothelial cells (ECs). Pyroptosis is a novel form of inflammatory caspase-1-dependent programmed cell death that is closely associated with the activation of the NOD-like receptor 3 (NLRP3) inflammasome. Recent studies have shown that sulforaphane (SFN) can inhibit pyroptosis, but the effects and underlying mechanisms by which SFN affects MGO-induced pyroptosis in endothelial cells have not been determined. Here, we found that SFN prevented MGO-induced pyroptosis by suppressing oxidative stress and inflammation in vitro and in vivo. Our results revealed that SFN dose-dependently prevented MGO-induced HUVEC pyroptosis, inhibited pyroptosis-associated biochemical changes, and attenuated MGO-induced morphological alterations in mitochondria. SFN pretreatment significantly suppressed MGO-induced ROS production and the inflammatory response by inhibiting the NLRP3 inflammasome (NLRP3, ASC, and caspase-1) signaling pathway by activating Nrf2/HO-1 signaling. Similar results were obtained in vivo, and we demonstrated that SFN prevented MGO-induced oxidative damage, inflammation and pyroptosis by reversing the MGO-induced downregulation of the NLRP3 signaling pathway through the upregulation of Nrf2. Additionally, an Nrf2 inhibitor (ML385) noticeably attenuated the protective effects of SFN on MGO-induced pyroptosis and ROS generation by inhibiting the Nrf2/HO-1 signaling pathway, and a ROS scavenger (NAC) and a permeability transition pore inhibitor (CsA) completely reversed these effects. Moreover, NLRP3 inhibitor (MCC950) and caspase-1 inhibitor (VX765) further reduced pyroptosis in endothelial cells that were pretreated with SFN. Collectively, these findings broaden our understanding of the mechanism by which SFN inhibits pyroptosis induced by MGO and suggests important implications for the potential use of SFN in the treatment of vascular diseases.
10.1007/s00018-024-05432-8
Resveratrol Ameliorates Deep Vein Thrombosis-Induced Inflammatory Response Through Inhibiting HIF-1α/NLRP3 Pathway.
Inflammation
Deep vein thrombosis (DVT) has become a prevalent and increasingly serious problem globally and resveratrol (Res) is a natural antitoxin that inhibits arterial thrombosis. To investigate the effect of Res on DVT and further explore its mechanism, thrombosis was monitored at different time points and the pathological changes occurring in the inferior vena cava (IVC) and lung tissue were observed in Sprague-Dawley rats. The protein expression of HIF-1α and NLRP3 in the IVC and lung tissue and the concentrations of D-dimer (D2D), prothrombin fragment 1 + 2 (F1 + 2), interleukin-1β (IL-1β), caspase-1, and tissue factor (TF) in the plasma were determined. After setting different doses of Res groups and using low-molecular-weight heparin (LMWH) as a positive control to determine the effective experimental dose of Res, rats were further divided into sham, DVT, HIF-1α inhibitor, Res, and HIF-1α inhibitor + Res groups. The above indicators were tested repeatedly. The DVT was formed on the 1st day of modeling. With the extension of time, DVT was gradually institutionalized and finally recanalized. Lesions in the IVC and lung tissue were effectively ameliorated, and thrombosis was significantly decreased in the LMWH or 60 mg/kg Res-treated groups. The levels of D2D, F1 + 2, IL-1β, caspase-1, TF, and the expression of HIF-1α and NLRP3 were significantly reduced in the HIF-1α inhibitor, Res, and HIF-1α inhibitor + Res groups. Res can ameliorate DVT in rats by inhibiting HIF-1α/NLRP3 pathway, which provides a novel therapeutic strategy for DVT treatment.
10.1007/s10753-022-01689-y