Protease-activated Receptor Research Articles

SCAR-6 elncRNA locus epigenetically regulates PROZ and modulates coagulation and vascular function

In this study, we characterize a novel lncRNA-producing gene locus that we name Syntenic Cardiovascular Conserved Region-Associated lncRNA-6 (scar-6) and functionally validate its role in coagulation and cardiovascular function. A 12-bp deletion of the scar-6 locus in zebrafish (scar-6gib007Δ12/Δ12) results in cranial hemorrhage and vascular permeability. Overexpression, knockdown and rescue with the scar-6 lncRNA modulates hemostasis in zebrafish. Molecular investigation reveals that the scar-6 lncRNA acts as an enhancer lncRNA (elncRNA), and controls the expression of prozb, an inhibitor of factor Xa, through an enhancer element in the scar-6 locus. The scar-6 locus suppresses loop formation between prozb and scar-6 sequences, which might be facilitated by the methylation of CpG islands via the prdm14-PRC2 complex whose binding to the locus might be stabilized by the scar-6 elncRNA transcript. Binding of prdm14 to the scar-6 locus is impaired in scar-6gib007Δ12/Δ12 zebrafish. Finally, activation of the PAR2 receptor in scar-6gib007Δ12/Δ12 zebrafish triggers NF-κB-mediated endothelial cell activation, leading to vascular dysfunction and hemorrhage. We present evidence that the scar-6 locus plays a role in regulating the expression of the coagulation cascade gene prozb and maintains vascular homeostasis.

Targeting PAR2-mediated inflammation in osteoarthritis: a comprehensive in vitro evaluation of oleocanthal’s potential as a functional food intervention for chondrocyte protection and anti-inflammatory effects

BackgroundOsteoarthritis (OA) is a prevalent degenerative joint disease characterized by chronic inflammation and progressive cartilage degradation, ultimately leading to joint dysfunction and disability. Oleocanthal (OC), a bioactive phenolic compound derived from extra virgin olive oil, has garnered significant attention due to its potent anti-inflammatory properties, which are comparable to those of non-steroidal anti-inflammatory drugs (NSAIDs). This study pioneers the investigation into the effects of OC on the Protease-Activated Receptor-2 (PAR-2) mediated inflammatory pathway in OA, aiming to validate its efficacy as a functional food-based therapeutic intervention.MethodsTo simulate cartilage tissue in vitro, human bone marrow-derived mesenchymal stem cells (BMSCs) were differentiated into chondrocytes. An inflammatory OA-like environment was induced in these chondrocytes using lipopolysaccharide (LPS) to mimic the pathological conditions of OA. The therapeutic effects of OC were evaluated by treating these inflamed chondrocytes with various concentrations of OC. The study focused on assessing key inflammatory markers, catabolic enzymes, and mitochondrial function to elucidate the protective mechanisms of OC. Mitochondrial function, specifically mitochondrial membrane potential (ΔΨm), was assessed using Rhodamine 123 staining, a fluorescent dye that selectively accumulates in active mitochondria. The integrity of ΔΨm serves as an indicator of mitochondrial and bioenergetic function. Additionally, Western blotting was employed to analyze protein expression levels, while real-time polymerase chain reaction (RT-PCR) was used to quantify gene expression of inflammatory cytokines and catabolic enzymes. Flow cytometry was utilized to measure cell viability and apoptosis, providing a comprehensive evaluation of OC’s therapeutic effects on chondrocytes.ResultsThe results demonstrated that OC significantly downregulated PAR-2 expression in a dose-dependent manner, leading to a substantial reduction in pro-inflammatory cytokines, including TNF-α, IL-1β, and MCP-1. Furthermore, OC attenuated the expression of catabolic markers such as SOX4 and ADAMTS5, which are critically involved in cartilage matrix degradation. Importantly, OC was found to preserve mitochondrial membrane potential (ΔΨm) in chondrocytes subjected to inflammatory stress, as evidenced by Rhodamine 123 staining, indicating a protective effect on cellular bioenergetics. Additionally, OC modulated the Receptor Activator of Nuclear Factor Kappa-Β Ligand (RANKL)/Receptor Activator of Nuclear Factor Kappa-Β (RANK) pathway, suggesting a broader therapeutic action against the multifactorial pathogenesis of OA.ConclusionsThis study is the first to elucidate the modulatory effects of OC on the PAR-2 mediated inflammatory pathway in OA, revealing its potential as a multifaceted therapeutic agent that not only mitigates inflammation but also protects cartilage integrity. The preservation of mitochondrial function and modulation of the RANKL/RANK pathway further underscores OC’s comprehensive therapeutic potential in counteracting the complex pathogenesis of OA. These findings position OC as a promising candidate for integration into nutritional interventions aimed at managing OA. However, further research is warranted to fully explore OC’s therapeutic potential across different stages of OA and its long-term effects in musculoskeletal disorders.

Integrative phosphoproteomic analyses reveal hemostatic-endothelial signaling interplay

BackgroundThe vascular endothelial cell (EC) monolayer plays a crucial part in maintaining hemostasis. An extensive array of G protein-coupled receptors allows ECs to dynamically act on key hemostatic stimuli such as thrombin and histamine. The impact of these individual stimuli on EC signal transduction has been the subject of various studies, but insight into discordant and concordant EC signaling between different G protein-coupled receptors remains limited. ObjectivesTo elucidate histamine and protease-activated receptor (PAR1-4) signaling cascades in ECs, discern overlapping and diverging regulation between these stimuli and their effect on the EC monolayer. MethodsWe employed stable isotope labeling by amino acids in cell culture mass spectrometry-based phosphoproteomics on in vitro cultured blood outgrowth ECs stimulated with histamine and different PAR1 to 4 peptides. We investigated key phosphosites through immuno(fluorescence) staining and determined effects on barrier function through transendothelial resistance assays. ResultsEC histamine activation initiated an extensive (kinase) signaling network (including MAPK3, STAT3, and CTNND1). PAR1 and PAR2 receptors induced highly similar signaling cascades, whereas PAR3 and PAR4 induced minimal phospho-regulation. Integration of all applied stimuli indicated uniquely activated proteins between both stimuli, as well as a general overlapping activation of cell junction and actin cytoskeletal proteins. ConclusionWe provide an integrative phosphoproteomic analysis of histamine and PAR agonists in the endothelium that highlights the endothelial response programs that are at the basis of regulating hemostasis.

Macrophage-Expressed Coagulation Factor VII Promotes Adverse Cardiac Remodeling.

Excess fibrotic remodeling causes cardiac dysfunction in ischemic heart disease, driven by MAP (mitogen-activated protein) kinase-dependent TGF-ß1 (transforming growth factor-ß1) activation by coagulation signaling of myeloid cells. How coagulation-inflammatory circuits can be specifically targeted to achieve beneficial macrophage reprogramming after myocardial infarction (MI) is not completely understood. Mice with permanent ligation of the left anterior descending artery were used to model nonreperfused MI and analyzed by single-cell RNA sequencing, protein expression changes, confocal microscopy, and longitudinal monitoring of recovery. We probed the role of the tissue factor (TF)-FVIIa (activated factor VII)-integrin ß1-PAR2 (protease-activated receptor 2) signaling complex by utilizing genetic mouse models and pharmacological intervention. Cleavage-insensitive PAR2R38E and myeloid cell integrin ß1-deficient mice had improved cardiac function after MI compared with controls. Proximity ligation assays of monocytic cells demonstrated that colocalization of FVIIa with integrin ß1 was diminished in monocyte/macrophage FVII-deficient mice after MI. Compared with controls, F7fl/fl CX3CR1 (CX3C motif chemokine receptor 1)Cre mice showed reduced TGF-ß1 and MAP kinase activation, as well as cardiac dysfunction after MI, despite unaltered overall recruitment of myeloid cells. Single-cell mRNA sequencing of CD45 (cluster of differentiation 45)+ cells 3 and 7 days after MI uncovered a trajectory from recruited monocytes to inflammatory TF+/TREM (triggered receptor expressed on myeloid cells) 1+ macrophages requiring F7. As early as 7 days after MI, macrophage F7 deletion led to an expansion of reparative Olfml 3 (olfactomedin-like protein 3)+ macrophages and, conversely, to a reduction of TF+/TREM1+ macrophages, which were also reduced in PAR2R38E mice. Short-term treatment from days 1 to 5 after nonreperfused MI with a monoclonal antibody inhibiting the macrophage TF-FVIIa-PAR2 signaling complex without anticoagulant activity improved cardiac dysfunction, decreased excess fibrosis, attenuated vascular endothelial dysfunction, and increased survival 28 days after MI. Extravascular TF-FVIIa-PAR2 complex signaling drives inflammatory macrophage polarization in ischemic heart disease. Targeting this signaling complex for specific therapeutic macrophage reprogramming following MI attenuates cardiac fibrosis and improves cardiovascular function.

Visualization of Neutrophil Extracellular Traps in Mesenteric Venules After Mesenteric Ischemia-Reperfusion Injury via Intravital Microscopy.

Intestinal ischemia-reperfusion (I/R) injury is an acute condition characterized by tissue damage resulting from restricted blood flow to the mesenteric vessels, leading to both local and systemic pathologies with a poor prognosis. Both ischemia and reperfusion trigger a series of cellular and molecular responses, with inflammatory cells serving as key regulators of the pathology. These interactions with the ischemic endothelium are mediated by multiple adhesion receptors. Several animal models have been established to mimic this pathology and investigate the involved molecular pathways. In this study, a microsurgical model of I/R injury is combined with intravital microscopy to visualize leukocyte rolling, adhesion, and neutrophil extracellular trap (NET) formation. This model is applied to transgenic mice deficient in endothelial PAR1 (F2r) to assess the impact of PAR1 on leukocyte rolling and NET formation 1 h after ischemia and immediately following reperfusion. In vivo, Acridine Orange leukocyte staining was employed, and NETs were visualized using a nucleic acid stain. Interestingly, reduced leukocyte adhesion and NET formation were observed in mice lacking the endothelial PAR1 receptor. This model enables the in vivo analysis of key regulators involved in I/R injury.

Differential Effect of Omega-3 Fatty Acids on Platelet Inhibition by Antiplatelet Drugs In Vitro.

The omega-3 polyunsaturated fatty acids (PUFAs) Docosahexaenoic acid (DHA) and Eicosapentaenoic acid (EPA) exert multiple cardioprotective effects, influencing inflammation, platelet activation, endothelial function and lipid metabolism, besides their well-established triglyceride lowering properties. It is not uncommon for omega-3 PUFAs to be prescribed for hypertriglyceridemia, alongside antiplatelet therapy in cardiovascular disease (CVD) patients. In this regard, we studied the effect of EPA and DHA, in combination with antiplatelet drugs, in platelet aggregation and P-selectin and αIIbβ3 membrane expression. The antiplatelet drugs aspirin and triflusal, inhibitors of cyclooxygenase-1 (COX-1); ticagrelor, an inhibitor of the receptor P2Y12; vorapaxar, an inhibitor of the PAR-1 receptor, were combined with DHA or EPA and evaluated against in vitro platelet aggregation induced by agonists arachidonic acid (AA), adenosine diphosphate (ADP) and TRAP-6. We further investigated procaspase-activating compound 1 (PAC-1) binding and P-selectin membrane expression in platelets stimulated with ADP and TRAP-6. Both DHA and EPA displayed a dose-dependent inhibitory effect on platelet aggregation induced by AA, ADP and TRAP-6. In platelet aggregation induced by AA, DHA significantly improved acetylsalicylic acid (ASA) and triflusal's inhibitory activity, while EPA enhanced the inhibitory effect of ASA. In combination with EPA, ASA and ticagrelor expressed an increased inhibitory effect towards ADP-induced platelet activation. Both fatty acids could not improve the inhibitory effect of vorapaxar on AA- and ADP-induced platelet aggregation. In the presence of EPA, all antiplatelet drugs displayed a stronger inhibitory effect towards TRAP-6-induced platelet activation. Both omega-3 PUFAs inhibited the membrane expression of αIIbβ3, though they had no effect on P-selectin expression induced by ADP or TRAP-6. The antiplatelet drugs exhibited heterogeneity regarding their effect on P-selectin and αIIbβ3 membrane expression, while both omega-3 PUFAs inhibited the membrane expression of αIIbβ3, though had no effect on P-selectin expression induced by ADP or TRAP-6. The combinatory effect of DHA and EPA with the antiplatelet drugs did not result in enhanced inhibitory activity compared to the sum of the individual effects of each component.

Update on protease-activated receptor 2 in inflammatory and autoimmune dermatological diseases.

Protease-activated receptor 2 (PAR2) is a cell-surface receptor expressed in various cell types, including keratinocytes, neurons, immune and inflammatory cells. Activation of PAR2, whether via its canonical or biased pathways, triggers a series of signaling cascades that mediate numerous functions. This review aims to highlight the emerging roles and interactions of PAR2 in different skin cells. It specifically summarizes the latest insights into the roles of PAR2 in skin conditions such as atopic dermatitis (AD), psoriasis, vitiligo and melasma. It also considers these roles from the perspective of the cutaneous microenvironment in relation to other inflammatory and autoimmune dermatological disorders. Additionally, the review explores PAR2's involvement in associated comorbidities from both cutaneous and extracutaneous diseases. Therefore, PAR2 may serve as a key target for interactions among various cells within the local skin environment.

Thrombin receptor PAR4 cross-activates the tyrosine kinase c-met in atrial cardiomyocytes.

Thrombin supports coagulation-independent inflammation via protease-activated receptors (PAR). PAR4 is specifically increased in obese human atria, correlating with NLRP3 inflammasome activation. PAR4-mediated NLRP3 inflammasome activation in atrial cardiomyocytes is not known, nor have signaling partners been identified. Thrombin transactivates the hepatocyte growth factor receptor in some cancer cells, so we examined PAR4/c-met cross-talk in atrial cardiomyocytes and its possible significance in obesity. Cardiomyocytes from right atrial appendages (RAA) of obese patients expressed more PAR1 and PAR4 compared to non-obese. In HL-1 atrial cardiomyocytes, thrombin induced caspase-1 auto-activation and IL-1β maturation; IL-1β secretion was evoked by PAR4-activating peptide (AP), but not PAR1-AP. PAR4-AP additionally increased phosphorylated CaMKII-Thr287, mTOR-Ser2481, and Akt-Ser473 while suppressing AMPK-Thr172 phosphorylation. Total kinase levels were largely unaltered. PAR4AP rapidly increased phosphorylated c-met in HL-1 cells and over time also transcriptionally upregulated c-met. The c-met inhibitor SGX-523 abrogated the effects of PAR4-AP on CaMKII/AKT/mTOR phosphorylation but did not affect PAR4-stimulated IL-1β production. Obese human RAA contained more IL-1β, phospho-c-met, and phospho-mTOR than non-obese RAA; CamKII phosphorylation was not modified. Atria from high-fat diet (HFD) versus chow-fed mice also contained more IL-1β, together with higher myeloperoxidase activity, Acta2 mRNA total and phosphorylated c-met; these increases were blunted in PAR4-/- HFD-fed mice. Thrombin cross-activates c-met via PAR4 in atrial cardiomyocytes. Transactivated c-met contributes partially to PAR4-mediated signaling, but NLRP3 inflammasome activation appears to be largely independent of c-met. Abundance of PAR4 and activated c-met increases with obesity, providing therapeutic targets for management of adiposity-driven AF.

The Role of Tissue Factor In Signaling Pathways of Pathological Conditions and Angiogenesis.

Tissue factor (TF) is an integral transmembrane protein associated with the extrinsic coagulation pathway. TF gene expression is regulated in response to inflammatory cytokines, bacterial lipopolysaccharides, and mechanical injuries. TF activity may be affected by phosphorylation of its cytoplasmic domain and alternative splicing. TF acts as the primary initiator of physiological hemostasis, which prevents local bleeding at the injury site. However, aberrant expression of TF, accompanied by the severity of diseases and infections under various pathological conditions, triggers multiple signaling pathways that support thrombosis, angiogenesis, inflammation, and metastasis. Protease-activated receptors (PARs) are central in the downstream signaling pathways of TF. In this study, we have reviewed the TF signaling pathways in different pathological conditions, such as wound injury, asthma, cardiovascular diseases (CVDs), viral infections, cancer and pathological angiogenesis. Angiogenic activities of TF are critical in the repair of wound injuries and aggressive behavior of tumors, which are mainly performed by the actions of vascular endothelial growth factor (VEGF) and hypoxia-inducible factor-1 (HIF1-α). Pro-inflammatory effects of TF have been reported in asthma, CVDs and viral infections, including COVID-19, which result in tissue hypertrophy, inflammation, and thrombosis. TF-FVII induces angiogenesis via clotting-dependent and -independent mechanisms. Clottingdependent angiogenesis is induced via the generation of thrombin and cross-linked fibrin network, which facilitate vessel infiltration and also act as a reservoir for endothelial cells (ECs) growth factors. Expression of TF in tumor cells and ECs triggers clotting-independent angiogenesis through induction of VEGF, urokinase-type plasminogen activator (uPAR), early growth response 1 (EGR1), IL8, and cysteine-rich angiogenic inducer 61 (Cyr61).

Abstract 43: PAR1 Inhibition with Vorapaxar Enhances Renal Function and Modulates Inflammation Mediators in Diabetic Nephropathy

Diabetic nephropathy (DN) is the leading cause of end-stage renal failure, and hypertension is a comorbidity in about 90% of all cases. Characteristics of pathological changes of DN include severe albuminuria, renal hyperfiltration, glomerular basement membrane thickening, and glomerulosclerosis. Identifying the molecular mechanisms underlying DN is critical for developing more effective therapeutic approaches. Thus, interventions targeting podocytes have become a major focus for diseases like DN due to their crucial role in regulating glomerular permeability and maintaining glomerular structures. Our previous studies have demonstrated that serine proteases and Protease-activated receptor 1 (PAR1) are involved in developing DN through TRPC6-mediated calcium signaling in podocytes. Utilizing a PAR1 heterozygous knockout rat model with a type 2 diabetic background, we showed that reducing PAR1 expression decreases microalbuminuria and glomerular damage. This study aims to define the effects and potential mechanisms of pharmacological blockade of PAR1 on the development of DN. To achieve this, 16-week-old T2DN rats were treated with a potent and selective PAR1 antagonist (vorapaxar (SCH 530348), 10 mg/kg of diet) for 16 weeks to determine the effect of PAR1 inhibition. We found improvements in glomerular filtration rate (GFR) in treated animals compared to control after 16 weeks of treatment (0.8 ± 0.2 vs 1.2 ± 0.2 mL/min/100g b.w., p=0.029, N≥4). We observed a trend toward differences in basal intracellular calcium concentration in podocytes between the control and treated groups, although these changes were not statistically significant (202 ± 95 vs 152 ± 75 nM, p=0.19, n≥10, N=3). Additionally, inhibition of PAR1 caused modulation in the inflammation-resolution mediators in the kidney with significant upregulation of Maresins Conjugates in Tissue Regeneration (MCTR1) and downregulation of hydroxy-docosahexaenoic acids (7-,14-,17-HDHA). Changes in GFR may be linked to reduced glomerular damage, indicating that the structural integrity of the glomeruli is better maintained in treated animals. Alterations in the lipidomic profile could indicate a reduction in the inflammatory processes typically associated with DN. Overall, our data indicates that vorapaxar improves glomerular filtration rate and modulates lipid mediators in a type 2 diabetic rat model. This suggests that targeting PAR1 could be a promising therapeutic strategy for diabetic nephropathy.

Liposomal fusion of plant-based extracellular vesicles to enhance skin anti-inflammation

This study reports a hybrid extracellular vesicle (EV)/liposome system developed through vesicular membrane fusion between anti-inflammatory Hydrangea macrophylla leaf EVs (HML-EVs) and terpinen-4-ol/azelamide monoethanolamine-loaded liposomes. Successful liposomal fusion between HML-EVs and terpinen-4-ol/azelamide monoethanolamine-loaded-liposome membranes, confirmed using the two-color fluorescence resonance energy transfer mechanism, enabled the scaled production of a stable hybrid HML-EV/liposome dispersion at a length scale of 100–200 nm. We show that the hybrid EV/liposome significantly inhibits apoptosis by nitric oxide and tumor necrosis factor-α production. Furthermore, HML-EV/liposomes can inhibit the synthesis of protease-activated receptor-2, interleukin-6, c-Jun, and thymic stromal lymphopoietin, thus exhibiting anti-inflammatory performance. These findings highlight that the hybrid EV/liposome system established in this study alleviates skin irritation and improves skin anti-inflammation.

Protease activated receptor-1 regulates mixed lineage kinase-3 to drive triple-negative breast cancer tumorigenesis

Triple-negative breast cancer (TNBC) is difficult to treat breast cancer subtype due to lack or insignificant expressions of targetable estrogen receptor (ER) and human epidermal growth factor receptor 2 (HER2). Therefore, finding a targetable protein or signaling pathway in TNBC would impact patient care. Here, we report that a member of the Mixed Lineage Kinase (MLK) family, MLK3, is an effector of G-protein-coupled protease-activated receptors 1 (PAR1) and targeting MLK3 by a small-molecule inhibitor prevented PAR1-mediated TNBC tumorigenesis. In silico and immunohistochemistry analysis of human breast tumors showed overexpression of PAR1 and MLK3 in TNBC tumors. Treating α-thrombin and PAR1 agonist increased MLK3 and JNK activities and induced cell migration in TNBC cells. The PAR1 positive/high (PAR1+/hi) population of TNBC cells showed aggressive tumor phenotype with increased MLK3 signaling. Moreover, combined inhibition of the PAR1 and MLK3 mitigated the TNBC tumor burden in preclinical TNBC models. Our data suggests that activation of the PAR1-MLK3 axis promotes TNBC tumorigenesis. Therefore, combinatorial therapy targeting MLK3 and PAR1 could effectively reduce TNBC tumor burden.

Long-term GABA Supplementation Regulates Diabetic Gastroenteropathy through GABA Receptor/trypsin-1/PARs/Akt/COX-2 Axis.

Molecular alterations of diabetic gastroenteropathy are poorly identified. This study investigates the effects of prolonged GABA supplementation on key protein expression levels of trypsin-1, PAR-1, PAR-2, PAR-3, PI3K, Akt, COX-2, GABAA, and GABAB receptors in the gastric tissue of type 2 diabetic rats (T2DM). To induce T2DM, a 3-month high-fat diet and 35 mg/kg of streptozotocin was used. Twenty-four male Wistar rats were divided into 4 groups: (1) control, (2) T2DM, (3) insulin-treated (2.5 U/kg), and (4)GABA-treated (1.5 g/kg GABA). Blood glucose was measured weekly. The protein expressions were assessed using western blotting. Histopathological changes were examined by H&E and Masson's staining. Diabetic rats show reduced NOS1 and elevated COX-2 and trypsin-1 protein expression levels in gastric tissue. Insulin and GABA therapy restored the NOS1 and COX-2 levels to control values. Insulin treatment increased PI3K, Akt, and p-Akt and, decreased trypsin-1, PAR-1, PAR-2, and PAR-3 levels in the diabetic rats. Levels of GABAA and GABAB receptors normalized following insulin and GABA therapy. H&E staining indicated an increase in mucin secretion following GABA treatment. These results suggest that GABA by acting on GABA receptors may regulate the trypsin-1/PARs/Akt/COX-2 pathway and thereby improve complications of diabetic gastroenteropathy.

DIFFERENCES IN PROTEASE ACTIVATED RECEPTOR-1 AND THROMBINE LEVELS IN PREECLAMPSIA AND NORMAL PREGNANCY

Introduction: Preeclampsia is a condition caused by alterations in endothelial function during pregnancy. Changes in endothelial function result in an increase in coagulation and microvascular fibrin accumulation, which results in impaired placental perfusion. Thrombin, which converts fibrin to fibrinogen, as well as platelet activity, the fibrinolytic system, and anticoagulants, are all procoagulant circumstances in preeclampsia. Thrombin contributes to the pathogenesis of preeclampsia by increasing the expression of sFlt-1 thereby providing an antiangiogenic response. Protease Activated Receptor-1 (PAR-1) is a mediator of thrombin for coagulation and inflammation in preeclampsia. Inhibition of Protease Activated Receptor-1 expression in trophoblasts can enhance placental angiogenesis and vascular remodeling. Recently, only few studies have assessed the levels of Protease Activated Receptor -7 and thrombin in preeclampsia.Objective: To determine the difference in levels of Protease Activated Receptor-1 and thrombin in preeclampsia and normal pregnancyMethods: This study is observational with a cross-sectional comparative study design. Sampling was conducted from March 2020 to March 2021. A total of 66 patients were investigated, with 33 samples of preeclampsia and 33 samples of normal pregnancy. The independent sample T-test was used for statistical analysis.Results: The mean levels of Protease Activated Receptor-1 in the preeclampsia group were higher at 28.56 ± 7.68 ng/mL while normal pregnancy was 21.67 ± 6.92 ng/mL. The results of statistical tests showed that there was a significant difference in levels of Protease Activated Receptor-1 between the preeclampsia and normal pregnancy groups (p<0.05). The mean thrombin level in the preeclampsia group was higher at 72.23 ± 7.99 ng/mL, while in normal pregnancy it was 63.70 ± 8.92 ng/mL. The difference in thrombin levels between the preeclampsia and normal pregnancy groups was statistically significant (p<0.05).Conclusion: Preeclampsia was associated with greater levels of Protease Activated Receptor-1 and thrombin than normal pregnancy. There was a significant difference in the mean levels of Protease Activated Receptor-1 and thrombin between preeclampsia and normal pregnancy.Keywords: Thrombin, Protease Activated Receptor-1(PAR-1), Preeclampsia

The biochemistry of melanogenesis: an insight into the function and mechanism of melanogenesis-related proteins.

Melanin is an amino acid derivative produced by melanocyte through a series of enzymatic reactions using tyrosinase as substrate. Human skin and hair color is also closely related to melanin, so understanding the mechanisms and proteins that produce melanin is very important. There are many proteins involved in the process of melanin expression, For example, proteins involved in melanin formation such as p53, HNF-1α (Hepatocyte nuclear factor 1α), SOX10 (Sry-related HMg-Box gene 10) and pax3 (paired box gene 3), MC1R(Melanocortin 1 Receptor), MITF (Microphthalmia-associated transcription factor), TYR (tyrosinase), TYRP1 (tyrosinase-related protein-1), TYRP2 (tyrosinase-related protein-2), and can be regulated by changing their content to control the production rate of melanin. Others, such as OA1 (ocular albinism type 1), Par-2 (protease-activated receptor 2) and Mlph (Melanophilin), have been found to control the transfer rate of melanosomes from melanocytes to keratinocytes, and regulate the amount of human epidermal melanin to control the depth of human skin color. In addition to the above proteins, there are other protein families also involved in the process of melanin expression, such as BLOC, Rab and Rho. This article reviews the origin of melanocytes, the related proteins affecting melanin and the basic causes of related gene mutations. In addition, we also summarized the active ingredients of 5 popular whitening cosmetics and their mechanisms of action.

Mechanisms and Therapeutic Targets of Staphylococcus Aureus - Induced Itch in Atopic Dermatitis

Atopic dermatitis is a chronic skin disorder marked by inflammation, erythema, and pruritus, often beginning in childhood and characterized by exacerbations and remissions. Its etiology involves genetic, immunological, and environmental factors. Staphylococcus aureus, a common pathogen, exacerbates atopic dermatitis by producing toxins and enzymes such as the serine protease V8, which activates the protease-activated receptor 1 (PAR1) on sensory neurons, inducing itch. This mechanism highlights potential therapeutic targets for alleviating pruritus. Research into the physiological pathways of itch, including the role of PAR1 and other protease-activated receptors, reveals promising strategies for treatment. PAR1 antagonists could be repurposed to treat chronic itch, providing new therapeutic avenues. PAR1 and other protease-activated receptors, is crucial in developing new treatment strategies. Anti-pruritic therapies targeting these pathways, such as PAR1 antagonists, show promise in mitigating itch symptoms. Moreover, existing drugs that inhibit PAR1 could be repurposed for treating chronic itch, providing a new avenue for relief in patients with atopic dermatitis. Additionally, the evolutionary role of itch induced by microorganisms suggests pathogens may exploit neural reflexes to enhance their spread. Advancements in understanding the mechanisms behind S. aureus-induced itch and the physiological pathways involved offer promising new directions for therapeutic intervention. Advancements in understanding the mechanisms of Staphylococcal aureus-induced itch and associated physiological pathways offer promising directions for therapeutic intervention, potentially improving management and treatment outcomes for patients with atopic dermatitis and other pruritic conditions.

Beyond Anticoagulation: A Comprehensive Review of Non-Vitamin K Oral Anticoagulants (NOACs) in Inflammation and Protease-Activated Receptor Signaling.

Non-vitamin K oral anticoagulants (NOACs) have revolutionized anticoagulant therapy, offering improved safety and efficacy over traditional agents like warfarin. This review comprehensively examines the dual roles of NOACs-apixaban, rivaroxaban, edoxaban, and dabigatran-not only as anticoagulants, but also as modulators of inflammation via protease-activated receptor (PAR) signaling. We highlight the unique pharmacotherapeutic properties of each NOAC, supported by key clinical trials demonstrating their effectiveness in preventing thromboembolic events. Beyond their established anticoagulant roles, emerging research suggests that NOACs influence inflammation through PAR signaling pathways, implicating factors such as factor Xa (FXa) and thrombin in the modulation of inflammatory responses. This review synthesizes current evidence on the anti-inflammatory potential of NOACs, exploring their impact on inflammatory markers and conditions like atherosclerosis and diabetes. By delineating the mechanisms by which NOACs mediate anti-inflammatory effects, this work aims to expand their therapeutic utility, offering new perspectives for managing inflammatory diseases. Our findings underscore the broader clinical implications of NOACs, advocating for their consideration in therapeutic strategies aimed at addressing inflammation-related pathologies. This comprehensive synthesis not only enhances understanding of NOACs' multifaceted roles, but also paves the way for future research and clinical applications in inflammation and cardiovascular health.

Attenuation of Atherosclerosis with PAR4 Deficiency: Differential Platelet Outcomes in apoE -/- vs. Ldlr -/- Mice.

Cardiovascular disease (CVD) is a significant burden globally and, despite current therapeutics, remains the leading cause of death. Platelet inhibitors are of interest in CVD treatment to reduce thrombus formation post-plaque rupture as well their contribution to inflammation throughout the progression of atherosclerosis. Protease activated receptor 4 (PAR4) is a receptor highly expressed by platelets, strongly activated by thrombin, and plays a vital role in platelet activation and aggregation. However, the role of PAR4. Mice on a low-density lipoprotein receptor-deficient ( Ldlr -/- ) background were bred with Par4 deficient ( Par4 -/- ) mice to create Ldlr -/- /Par4 +/+ and Ldlr -/- /Par4 -/- cousin lines. Mice were fed high fat (42%) and cholesterol (0.2%) 'Western' diet for 12 weeks for all studies. Bone marrow transplant (BMT) studies were conducted by irradiating Ldlr -/- /Par4 +/+ and Ldlr -/- /Par4 -/- mice with 550 rads (2x, 4 hours apart) and then repopulated with Par4 +/+ or Par4 -/- bone marrow. To determine if the effects of thrombin were mediated solely by PAR4, the thrombin inhibitor dabigatran was added to the 'Western' diet. Ldlr -/- /Par4 -/- given dabigatran did not further decrease their atherosclerotic burden. Differences between apolipoprotein E deficient ( apoE -/- ) and Ldlr -/- platelets were assessed for changes in reactivity. We observed higher PAR4 abundance in arteries with atherosclerosis in human and mice versus healthy controls. PAR4 deficiency attenuated atherosclerosis in the aortic sinus and root versus proficient controls. BMT studies demonstrated this effect was due to hematopoietic cells, most likely platelets. PAR4 appeared to be acting independent of PAR1, as there werer no changes with addition of dabigatran to PAR4 deficient mice. apoE -/- platelets are hyperreactive compared to Ldlr -/- platelets. Hematopoietic-derived PAR4, most likely platelets, plays a vital role in the development and progression of atherosclerosis. Specific targeting of PAR4 may be a potential therapeutic target for CVD. Deficiency of protease-activated receptor 4 attenuates the development of diet-induced atherosclerosis in a Ldlr -/- mouse model. PAR4 deficiency in hematopoietic cells is atheroprotective. PAR4 deficiency accounts for the majority of thrombin-induced atherosclerosis in a Ldlr -/- mouse model. The examination of platelet-specific proteins and platelet activation should be carefully considered before using the apoE -/- or Ldlr -/- mouse models of atherosclerosis.

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

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.

FXa-mediated PAR-2 promotes the efficacy of immunotherapy for hepatocellular carcinoma through immune escape and anoikis resistance by inducing PD-L1 transcription

BackgroundThe high metastasis rate is one of the main reasons for the poor prognosis of patients with hepatocellular carcinoma (HCC). Coagulation factor Xa (FXa) and its receptor proteinase-activated receptor-2 (PAR-2)...