RESUMEN

Background: Chronic thromboembolic pulmonary hypertension (CTEPH) is a serious pulmonary vascular disease characterized by residual thrombi in the pulmonary arteries and distal pulmonary microvascular remodeling. The pathogenesis of CTEPH remains unclear, but many factors such as inflammation, immunity, coagulation and angiogenesis may be involved. Monocytes are important immune cells that can differentiate into macrophages and dendritic cells and play an important role in thrombus formation. However, the distribution, gene expression profile and differentiation trajectory of monocyte subsets in CTEPH patients have not been systematically studied. This study aims to reveal the characteristics and functions of monocytes in CTEPH patients using single-cell sequencing technology, and to provide new insights for the diagnosis and treatment of CTEPH. Methods: Single-cell RNA sequencing (scRNA-seq) were performed to analyze the transcriptomic features of peripheral blood mononuclear cells (PBMCs) from healthy controls, CTEPH patients and the tissues from CTEPH patients after the pulmonary endarterectomy (PEA). We established a CTEPH rat model with chronic pulmonary embolism caused by repeated injection of autologous thrombi through a central venous catheter, and used flow cytometry to detect the proportion changes of monocyte subsets in CTEPH patients and CTEPH rat model. We also observed the infiltration degree of macrophage subsets in thrombus tissue and their differentiation relationship with peripheral blood monocyte subsets by immunofluorescence staining. Results: The results showed that the monocyte subsets in peripheral blood of CTEPH patients changed significantly, especially the proportion of CD16+ monocyte subset increased. This monocyte subset had unique functional features at the transcriptomic level, involving processes such as cell adhesion, T cell activation, coagulation response and platelet activation, which may play an important role in pulmonary artery thrombus formation and pulmonary artery intimal remodeling. In addition, we also found that the macrophage subsets in pulmonary endarterectomy tissue of CTEPH patients showed pro-inflammatory and lipid metabolism reprogramming features, which may be related to the persistence and insolubility of pulmonary artery thrombi and the development of pulmonary hypertension. Finally, we also observed that CD16+ monocyte subset in peripheral blood of CTEPH patients may be recruited to pulmonary artery intimal tissue and differentiate into macrophage subset with high expression of IL-1ß, participating in disease progression. Conclusion: CD16+ monocytes subset had significant gene expression changes in CTEPH patients, related to platelet activation, coagulation response and inflammatory response. And we also found that these cells could migrate to the thrombus and differentiate into macrophages with high expression of IL-1ß involved in CTEPH disease progression. We believe that CD16+ monocytes are important participants in CTEPH and potential therapeutic targets.

Asunto(s)

Hipertensión Pulmonar , Monocitos , Embolia Pulmonar , Receptores de IgG , Análisis de la Célula Individual , Humanos , Hipertensión Pulmonar/etiología , Hipertensión Pulmonar/inmunología , Hipertensión Pulmonar/metabolismo , Monocitos/inmunología , Monocitos/metabolismo , Receptores de IgG/metabolismo , Embolia Pulmonar/inmunología , Embolia Pulmonar/metabolismo , Animales , Masculino , Enfermedad Crónica , Ratas , Femenino , Persona de Mediana Edad , Proteínas Ligadas a GPI/metabolismo , Modelos Animales de Enfermedad , Transcriptoma , Anciano , Arteria Pulmonar/metabolismo , Arteria Pulmonar/inmunología , Arteria Pulmonar/patología

RESUMEN

BACKGROUND: Pulmonary thromboembolism (PTE) is a cardiovascular emergency that can result in mortality. In the interleukin-33 (IL-33) /soluble suppression of tumorigenicity 2 (sST2) signaling pathway, increased sST2 is a cardiovascular risk factor. This study aimed to investigate the effectiveness of biomarkers in the IL-33/sST2 signaling pathway in determining PTE diagnosis, clinical severity, and mortality. METHOD: This study was conducted as a single-center, prospective, observational study. Patients admitted to the emergency department and diagnosed with PTE constituted the patient group (n = 112), and healthy volunteers with similar sociodemographic characteristics constituted the control group (n = 62). Biomarkers in the IL-33/sST2 signaling pathway were evaluated for diagnosis, clinical severity, and prognosis. RESULTS: IL-33 was lower in the patient group than in the control group (275.89 versus 403.35 pg/mL), while sST2 levels were higher in the patient group than in the control group (53.16 versus 11.78 ng/mL) (p < 0.001 and p = 0.001; respectively). The AUC of IL-33 to diagnose PTE was 0.656 (95 % CI: 0.580-0.726). The optimal IL-33 cut-off point to diagnose PTE was ≤304.11 pg/mL (56.2 % sensitivity, 79 % specificity). The AUC of sST2 to diagnose PTE was 0.818 (95 % CI: 0.752-0.872). The optimal sST2 cut-off point to diagnose PTE was >14.48 ng/mL (83 % sensitivity, 71 % specificity). IL-33 levels were lower in patients with mortality (169.85 versus 332.04 pg/mL) compared to patients without mortality, whereas sST2 levels were higher in patients with mortality (118.32 versus 28.07 ng/mL) compared to patients without mortality (p > 0.001 for both). The AUC of IL-33 to predict the mortality of PTE was 0.801 (95 % CI: 0.715-0.870). The optimal IL-33 cut-off point to predict the mortality of PTE was ≤212.05 pg/mL (75 % sensitivity, 79.5 % specificity). The AUC of sST2 to predict the mortality of PTE was 0.824 (95 % CI: 0.740-0.889). The optimal sST2 cut-off point to predict the mortality of PTE was >81 ng/mL (95.8 % sensitivity, 78.4 % specificity). CONCLUSION: In the IL-33/ST2 signaling pathway, decreased IL-33 and increased sST2 are valuable biomarkers for diagnosis and prediction of mortality in patients with PTE.

Asunto(s)

Biomarcadores , Proteína 1 Similar al Receptor de Interleucina-1 , Interleucina-33 , Embolia Pulmonar , Transducción de Señal , Humanos , Interleucina-33/sangre , Interleucina-33/metabolismo , Embolia Pulmonar/mortalidad , Embolia Pulmonar/diagnóstico , Embolia Pulmonar/metabolismo , Embolia Pulmonar/sangre , Proteína 1 Similar al Receptor de Interleucina-1/sangre , Proteína 1 Similar al Receptor de Interleucina-1/metabolismo , Masculino , Femenino , Persona de Mediana Edad , Biomarcadores/sangre , Biomarcadores/metabolismo , Estudios Prospectivos , Anciano , Adulto , Pronóstico , Curva ROC

RESUMEN

AIMS: To detect the expression of autophagy components, p38 MAPK (p38) and phosphorylated forkhead box transcription factor O-1 (pFoxO1) in pulmonary vascular endothelial cells of chronic thromboembolic pulmonary hypertension (CTEPH) rats and to investigate the possible mechanism through which tissue factor (TF) regulates autophagy. METHODS: Pulmonary artery endothelial cells (PAECs) were isolated from CTEPH (CTEPH group) and healthy rats (control group (ctrl group)) which were cocultured with TF at different time points including 12 h, 24 h, 48 h and doses including 0 nM,10 nM, 100 nM, 1µM, 10µM, 100µM and cocultured with TFPI at 48 h including 0 nM, 2.5 nM, 5 nM. The expression of forkhead box transcription factor O-1 (FoxO1), pFoxO1, p38, Beclin-1 and LC3B in PAECs was measured. Coimmunoprecipitation (co-IP) assays were used to detect the interaction between FoxO1 and LC3. RESULTS: The protein expression of p-FoxO1/FoxO1 was significantly lower in the CTEPH groups (cocultured with TF from 0 nM to 100 µM) than in the ctrl group at 12 h, 24 h, and 48 h (P < 0.05) and was significantly lower in the CTEPH groups (cocultured with TFPI from 0 nM to 5 nM) than in the ctrl group at 48 h (P < 0.05). The protein expression of p38 in the CTEPH groups treated with 0 nM, 10 nM, 100 nM or 1 µM TF for 48 h significantly increased than ctrl groups (P < 0.05) and was significantly increased in the CTEPH groups (cocultured with TFPI concentration from 0 nM to 5 nM) than in the ctrl group at 48 h (P < 0.05). The protein expression of Beclin1 at the same concentration (cocultured with TF from 0 nM to 100 µM) was significantly lower in the CTEPH groups than ctrl groups after 24 h and 48 h (P < 0.05) and was significantly decreased in the CTEPH groups (cocultured with TFPI concentration from 2.5 nM to 5 nM) than in the ctrl group at 48 h (P < 0.05). The protein expression of LC3-II/LC3-I at the same concentration (cocultured with TF 0 nM, 1 µM, 10 µM, and 100 µM) was significantly lower in the CTEPH than in the ctrl groups after 12 h (P < 0.05) and was significantly lower in the CTEPH groups (cocultured with TFPI concentration from 0 nM to 5 nM) than in the ctrl group at 48 h (P < 0.05). There were close interactions between FoxO1 and LC3 in the control and CTEPH groups at different doses and time points. CONCLUSION: The autophagic activity of PAECs from CTEPH rats was disrupted. TF, FoxO1 and p38 MAPK play key roles in the autophagic activity of PAECs. TF may regulate autophagic activity through the p38 MAPK-FoxO1 pathway.

Asunto(s)

Autofagia , Células Endoteliales , Hipertensión Pulmonar , Arteria Pulmonar , Ratas Sprague-Dawley , Tromboplastina , Proteínas Quinasas p38 Activadas por Mitógenos , Animales , Autofagia/fisiología , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismo , Arteria Pulmonar/metabolismo , Arteria Pulmonar/patología , Ratas , Masculino , Células Endoteliales/metabolismo , Células Cultivadas , Tromboplastina/metabolismo , Tromboplastina/biosíntesis , Hipertensión Pulmonar/metabolismo , Embolia Pulmonar/metabolismo , Embolia Pulmonar/patología , Enfermedad Crónica , Transducción de Señal/fisiología , Proteína Forkhead Box O1

RESUMEN

In this study, we examined the changes in the fibrinolytic system in a rabbit model of two acute pulmonary thromboembolisms (PTE). Fourteen healthy adult New Zealand white rabbits were divided into three groups: the single PTE group (five rabbits), the double PTE group (five rabbits), and the control group (four rabbits). A rabbit model of acute pulmonary embolism was established, and immunohistochemistry and polymerase chain reaction (PCR) were performed on tissue plasminogen activator (t-PA), plasminogen activator inhibitor-1 (PAI-1) in plasma, and pulmonary embolism tissue. Plasma results: 1) t-PA levels: one hour following the initial modeling, the levels of t-PA in the modeling groups were significantly lower than those in the control group (P<0.05). In addition, the t-PA levels in the double PTE group were found to be lower after the modeling, as compared to the pre-modeling period (P<0.05). One hour after the second modeling, the double PTE group had lower t-PA levels compared to the control group (P<0.05). However, t-PA rebounded two hours after modeling in the double PTE group. One week after the second modeling, the double PTE group had higher t-PA levels compared to the other two groups (P<0.05). 2) PAI-1 results: one hour after the initial modeling, PAI-1 levels in the two modeling groups were lower compared to the pre-modeling period and control groups (P<0.05). Two hours following modeling, PAI-1 levels in both modeling groups were lower compared to the control group (P<0.05). PAI-1 levels were lower in the double PTE group one and two hours after the second modeling compared to the other two groups and pre-modeling period (P<0.05). 3) The immunohistochemistry results: the expression of PAI-1 decreased in the two modeling groups, while t-PA expression increased compared to the control group. 4) PCR results: t-PA mRNA expression did not differ among the three groups. The PAI-1 mRNA expression was lower in the two PTE groups compared to the control group. We conclude that in the early stages of PTE, the local fibrinolytic activity of the thrombus is increased, which is favorable for thrombolysis. However, as the thrombus persists, the activity of the fibrinolytic system is inhibited, contributing to the development of chronic thromboembolic pulmonary hypertension.

Asunto(s)

Modelos Animales de Enfermedad , Fibrinólisis , Inhibidor 1 de Activador Plasminogénico , Embolia Pulmonar , Activador de Tejido Plasminógeno , Animales , Conejos , Embolia Pulmonar/metabolismo , Embolia Pulmonar/sangre , Embolia Pulmonar/patología , Inhibidor 1 de Activador Plasminogénico/metabolismo , Inhibidor 1 de Activador Plasminogénico/genética , Activador de Tejido Plasminógeno/metabolismo , Activador de Tejido Plasminógeno/genética , Masculino , ARN Mensajero/metabolismo , ARN Mensajero/genética , Pulmón/metabolismo

RESUMEN

Platelets assume a pivotal role in the pathogenesis of cardiovascular diseases (CVDs), emphasizing their significance in disease progression. Consequently, addressing CVDs necessitates a targeted approach focused on mitigating platelet activation. Eugenol, predominantly derived from clove oil, is recognized for its antibacterial, anticancer, and anti-inflammatory properties, rendering it a valuable medicinal agent. This investigation delves into the intricate mechanisms through which eugenol influences human platelets. At a low concentration of 2 µM, eugenol demonstrates inhibition of collagen and arachidonic acid (AA)-induced platelet aggregation. Notably, thrombin and U46619 remain unaffected by eugenol. Its modulatory effects extend to ATP release, P-selectin expression, and intracellular calcium levels ([Ca2+]i). Eugenol significantly inhibits various signaling cascades, including phospholipase Cγ2 (PLCγ2)/protein kinase C (PKC), phosphoinositide 3-kinase/Akt/glycogen synthase kinase-3ß, mitogen-activated protein kinases, and cytosolic phospholipase A2 (cPLA2)/thromboxane A2 (TxA2) formation induced by collagen. Eugenol selectively inhibited cPLA2/TxA2 phosphorylation induced by AA, not affecting p38 MAPK. In ADP-treated mice, eugenol reduced occluded lung vessels by platelet thrombi without extending bleeding time. In conclusion, eugenol exerts a potent inhibitory effect on platelet activation, achieved through the inhibition of the PLCγ2-PKC and cPLA2-TxA2 cascade, consequently suppressing platelet aggregation. These findings underscore the potential therapeutic applications of eugenol in CVDs.

Asunto(s)

Eugenol , Embolia Pulmonar , Humanos , Ratones , Animales , Eugenol/farmacología , Eugenol/uso terapéutico , Eugenol/metabolismo , Fosfolipasa C gamma/metabolismo , Fosfatidilinositol 3-Quinasas/metabolismo , Modelos Animales de Enfermedad , Activación Plaquetaria , Agregación Plaquetaria , Plaquetas/metabolismo , Fosforilación , Proteína Quinasa C/metabolismo , Tromboxano A2/metabolismo , Colágeno/metabolismo , Embolia Pulmonar/tratamiento farmacológico , Embolia Pulmonar/metabolismo , Fosfolipasas A2 Citosólicas/metabolismo

RESUMEN

BACKGROUND: Thrombin-activatable fibrinolysis inhibitor (TAFI) levels are positively correlated with the risk of thrombosis. The mechanism of how TAFI affects venous thromboembolism (VTE) remains uncertain. In addition, the role of sex on the risk of VTE has also been studied. However, their association also remains unclear. OBJECTIVES: To investigate how TAFI and/or sex affect venous thrombus stability and consequent pulmonary embolism (PE). METHODS: Ferric chloride-induced thrombi were formed within the femoral veins of male and female wild-type (WT) or TAFI-knockout (Cpb2-/-) mice. Thrombi were imaged over 2 hours using intravital videomicroscopy to quantify embolization and thrombus size over time. Lungs were examined by immunohistochemistry to quantify (a) emboli and (b) fibrin composition of these emboli. RESULTS: Embolization events in female mice were higher than in males (7.9-fold in WT and 3.1-fold in Cpb2-/- mice). Although the maximal thrombus sizes were not different across groups, Cpb2-/- mice had thrombi that were, on average, 24% smaller at the end of the 2-hour experiment than WT mice. Loss of TAFI led to a 4.0- and 2.8-fold increase in PE burden in males and females, respectively, while sex had no influence. Pulmonary emboli in Cpb2-/- mice had higher fibrin composition compared with WT mice. CONCLUSION: Female mice had less stable venous thrombi than male mice, suggesting a higher risk of PE in females with deep vein thrombosis. Mice lacking TAFI had more thrombus degradation and higher PE burden than WT mice. These results confirm the role of TAFI in venous thrombosis.

Asunto(s)

Carboxipeptidasa B2 , Embolia Pulmonar , Trombosis , Tromboembolia Venosa , Masculino , Femenino , Ratones , Animales , Carboxipeptidasa B2/genética , Modelos Animales de Enfermedad , Embolia Pulmonar/genética , Embolia Pulmonar/metabolismo , Fibrina , Fibrinólisis

RESUMEN

Here we developed a model of chronic thromboembolic pulmonary hypertension (CTEPH) using repeated intravenous administration of microencapsulated thrombi with a controlled rate of biodegradation. Autologous thrombi encapsulated in alginate microspheres with a diameter of 190±48 µm were intravenously injected to rats 8 times every 4 days. In the comparison group, nonmodified thrombi were injected. After 6 weeks, a significant increase in systolic pressure in the right ventricle, a decrease in exercise tolerance, and an increase in the index of vascular wall hypertrophy were revealed in the group receiving injections of microencapsulated thrombi in comparison with the group receiving nonmodified thrombi and healthy animals. Thus, the developed representative CTEPH model can be used to test promising pharmacological substances.

Asunto(s)

Hipertensión Pulmonar , Embolia Pulmonar , Animales , Ratas , Hipertensión Pulmonar/metabolismo , Arteria Pulmonar/metabolismo , Embolia Pulmonar/metabolismo , Enfermedad Crónica

RESUMEN

AIMS: Platelet activation plays a central role in arterial thrombosis. Platelets are activated by adhesive proteins (i.e., collagen) or soluble agonists (i.e., thrombin), the respective receptor-specific signaling cause inside-out signaling, leading to the binding of fibrinogen to integrin αIIbß3. This binding triggers outside-in signaling, resulting in platelet aggregation. Garcinol, a polyisoprenylated benzophenone, is extracted from the fruit rind of Garcinia indica. Although garcinol exhibits considerable bioactivities, few studies have investigated the effect of garcinol on platelet activation. MAIN METHODS: Aggregometry, immunoblotting, flow cytometer, confocal microscopic analysis, fibrin clot retraction, animal studies such as fluorescein-induced platelet plug formation in mesenteric microvessels, acute pulmonary thromboembolism, and tail bleeding time were performed in this study. KEY FINDINGS: This study indicates that garcinol inhibited platelet aggregation stimulated by collagen, thrombin, arachidonic acid, and U46619. Garcinol reduced integrin αIIbß3 inside-out signaling, including ATP release; cytosolic Ca2+ mobilization; P-selectin expression; and Syk, PLCγ2/PKC, PI3K/Akt/GSK3ß, MAPKs, and NF-κB activation stimulated by collagen. Garcinol directly inhibited integrin αIIbß3 activation by interfering with FITC-PAC-1 and FITC-triflavin by collagen. Additionally, garcinol affected integrin αIIbß3-mediated outside-in signaling, such as decreasing platelet adhesion and the single-platelet spreading area; suppressing integrin ß3, Src, FAK, and Syk phosphorylation on immobilized fibrinogen; and inhibiting thrombin-stimulated fibrin clot retraction. Garcinol substantially reduced mortality caused by pulmonary thromboembolism and prolonged the occlusion time of thrombotic platelet plug formation without extending bleeding time in mice. SIGNIFICANCE: This study identified that garcinol, a novel antithrombotic agent, acts as a naturally occurring integrin αIIbß3 inhibitor.

Asunto(s)

Embolia Pulmonar , Trombosis , Humanos , Ratones , Animales , Plaquetas/metabolismo , Complejo GPIIb-IIIa de Glicoproteína Plaquetaria/metabolismo , Fluoresceína-5-Isotiocianato/metabolismo , Fosfatidilinositol 3-Quinasas/metabolismo , Trombina/metabolismo , Activación Plaquetaria , Agregación Plaquetaria , Trombosis/metabolismo , Fosforilación , Colágeno/metabolismo , Fibrinógeno/metabolismo , Embolia Pulmonar/metabolismo

RESUMEN

Venous thromboembolism (VTE) comprising deep venous thrombosis and pulmonary embolism is a major cause of morbidity and mortality. Short-term immobility-related conditions are a major risk factor for the development of VTE. Paradoxically, long-term immobilized free-ranging hibernating brown bears and paralyzed spinal cord injury (SCI) patients are protected from VTE. We aimed to identify mechanisms of immobility-associated VTE protection in a cross-species approach. Mass spectrometry-based proteomics revealed an antithrombotic signature in platelets of hibernating brown bears with heat shock protein 47 (HSP47) as the most substantially reduced protein. HSP47 down-regulation or ablation attenuated immune cell activation and neutrophil extracellular trap formation, contributing to thromboprotection in bears, SCI patients, and mice. This cross-species conserved platelet signature may give rise to antithrombotic therapeutics and prognostic markers beyond immobility-associated VTE.

Asunto(s)

Plaquetas , Proteínas del Choque Térmico HSP47 , Hipocinesia , Traumatismos de la Médula Espinal , Ursidae , Tromboembolia Venosa , Animales , Humanos , Ratones , Fibrinolíticos/uso terapéutico , Embolia Pulmonar/tratamiento farmacológico , Embolia Pulmonar/etnología , Embolia Pulmonar/metabolismo , Factores de Riesgo , Traumatismos de la Médula Espinal/complicaciones , Ursidae/metabolismo , Tromboembolia Venosa/etiología , Tromboembolia Venosa/metabolismo , Hipocinesia/complicaciones , Proteínas del Choque Térmico HSP47/metabolismo , Plaquetas/metabolismo

RESUMEN

INTRODUCTION: Acute pulmonary embolism (APE) is a clinical syndrome of pulmonary circulation disorder caused by obstruction of the pulmonary artery or its branches. Histone deacetylase 6 (HDAC6) has been reported to play an important role in lung-related diseases. However, the functional role of HDAC6 in APE remains unclear. MATERIAL AND METHODS: Male Sprague Dawley rats were used. The APE model was constructed by inserting an intravenous cannula into the right femoral vein and injecting Sephadex G-50 microspheres (12 mg/kg; 300 µm in diameter). After 1 h, the control and APE rats were intraperitoneally injected with tubastatin A (TubA) (40 mg/kg, an inhibitor of HDAC6) and sampled at 24 h after modeling. H&E staining, arterial blood gas analysis, and wet/dry (W/D) weight ratio were used to evaluate the histopathological changes and pulmonary function in APE rats. ELISA, Western blot, and immunohistochemistry were used to explore the potential mechanism of HDAC6-mediated inflammation in APE. RESULTS: The results indicated that HDAC6 expression was significantly increased in lungs of APE rats. TubA treatment in vivo decreased HDAC6 expression in lung tissues. HDAC6 inhibition alleviated histopathological damage and pulmonary dysfunction, as evidenced by decreased PaO2/FiO2 ratio and W/D weight ratio in APE rats. Furthermore, HDAC6 inhibition alleviated APE-induced inflammatory response. Specifically, APE rats exhibited increased production of pro-inflammatory cytokines, including tumor necrosis factor-alpha (TNF-α), interleukin (IL)-1ß, IL-6, and IL-18, however, this increase was reversed by HDAC6 inhibition. Meanwhile, the activation of the NLRP3 inflammasome was also observed in lungs of APE rats, while HDAC6 inhibition blocked this activation. Mechanically, we demonstrated that HDAC6 inhibition blocked the activation of the protein kinase B (AKT)/extracellular signal-regulated protein kinase (ERK) signaling pathway, a classic pathway promoting inflammation. CONCLUSIONS: These findings demonstrate that the inhibition of HDAC6 may alleviate lung dysfunction and pathological injury resulting from APE by blocking the AKT/ERK signaling pathway, providing new theoretical fundamentals for APE therapy.

Asunto(s)

Hominidae , Embolia Pulmonar , Ratas , Masculino , Animales , Proteínas Proto-Oncogénicas c-akt/metabolismo , Histona Desacetilasa 6 , Ratas Sprague-Dawley , Embolia Pulmonar/tratamiento farmacológico , Embolia Pulmonar/metabolismo , Inflamación , Factor de Necrosis Tumoral alfa , Quinasas MAP Reguladas por Señal Extracelular , Hominidae/metabolismo

RESUMEN

BACKGROUND: Pulmonary embolism (PE) is a severe disease that usually originates from deep vein thrombosis (DVT) of the lower extremities. This study set out to investigate the changes in the transcriptome of the pulmonary artery (PA) in the course of the PE in the porcine model. METHODS: The study was performed on 11 male pigs: a thrombus was formed in each right femoral vein in six animals, and then was released to induce PE, the remaining five animals served as a control group. In the experimental animals total RNA was isolated from the PA where the blood clot lodged, and in the control group, from the corresponding PA segments. High-throughput RNA sequencing was used to analyse the global changes in the transcriptome of PA with induced PE (PA-E). RESULTS: Applied multistep bioinformatics revealed 473 differentially expressed genes (DEGs): 198 upregulated and 275 downregulated. Functional Gene Ontology annotated 347 DEGs into 27 biological processes, 324 to the 11 cellular components and 346 to the 2 molecular functions categories. In the signaling pathway analysis, KEGG 'protein processing in endoplasmic reticulum' was identified for the mRNAs modulated during PE. The same KEGG pathway was also exposed by 8 differentially alternative splicing genes. Within single nucleotide variants, the 61 allele-specific expression variants were localised in the vicinity of the genes that belong to the cellular components of the 'endoplasmic reticulum'. The discovered allele-specific genes were also classified as signatures of the cardiovascular system. CONCLUSIONS: The findings of this research provide the first thorough investigation of the changes in the gene expression profile of PA affected by an embolus. Evidence from this study suggests that the disturbed homeostasis in the biosynthesis of proteins in the endoplasmic reticulum plays a major role in the pathogenesis of PE.

Asunto(s)

Embolia Pulmonar , Transcriptoma , Masculino , Animales , Porcinos , Arteria Pulmonar/metabolismo , Perfilación de la Expresión Génica , Embolia Pulmonar/genética , Embolia Pulmonar/metabolismo , Transducción de Señal

RESUMEN

Acute pulmonary thromboembolism (APTE) has become a non-negligible clinical concern due to its high mortality and complex symptoms. Early diagnosis and prognostic assessment of APTE are of great significance for the long-term benefits of patients, especially elderly patients. Elderly patients with pulmonary embolism (n = 250) who presented to our hospital from January 2018 to July 2021 were recruited into this study. In addition, 50 healthy elderly people with no history of allergies were selected as the control group. An enzyme-linked immunosorbent assay (ELISA) method was used to determine concentrations of D-dimer and signal peptide-CUB-EGF domain-containing protein-1 (SCUBE1) in their plasma. Right ventricular volume contraction time (ICT), ejection time (ET), and isovolumic relaxation time (IRT) were determined by Doppler ultrasound. Right ventricular Tei index was calculated as (ICT + IRT)/ET. High plasma D-dimer, plasma SCUBE1, and right ventricular Tei index are risk factors for poor prognosis in APTE patients after treatment. Plasma D-dimer, plasma SCUBE1, and right ventricular Tei index have predictive value for poor prognosis in APTE patients. Their combined detection (0.256*DD +0.04*SCUBE1 + 10.188*Tei) can improve the sensitivity and specificity of prediction. There is a predictive value of combined plasma D-dimer, SCUBE1, and right ventricular Tei index for the prognosis of elderly patients with APTE.

Asunto(s)

Productos de Degradación de Fibrina-Fibrinógeno , Embolia Pulmonar , Anciano , Humanos , Enfermedad Aguda , Proteínas de Unión al Calcio/química , Productos de Degradación de Fibrina-Fibrinógeno/química , Pronóstico , Embolia Pulmonar/sangre , Embolia Pulmonar/diagnóstico , Embolia Pulmonar/metabolismo

RESUMEN

Chronic thromboembolic pulmonary hypertension (CTEPH) and pulmonary arterial hypertension (PAH) are two forms of pulmonary hypertension (PH) characterized by obstructive vasculopathy. Endothelial dysfunction along with metabolic changes towards increased glycolysis are important in PAH pathophysiology. Less is known about such abnormalities in endothelial cells (ECs) from CTEPH patients. This study provides a systematic metabolic comparison of ECs derived from CTEPH and PAH patients. Metabolic gene expression was studied using qPCR in cultured CTEPH-EC and PAH-EC. Western blot analyses were done for HK2, LDHA, PDHA1, PDK and G6PD. Basal viability of CTEPH-EC and PAH-EC with the incubation with metabolic inhibitors was measured using colorimetric viability assays. Human pulmonary artery endothelial cells (HPAEC) were used as healthy controls. Whereas PAH-EC showed significant higher mRNA levels of GLUT1, HK2, LDHA, PDHA1 and GLUD1 metabolic enzymes compared to HPAEC, CTEPH-EC did not. Oxidative phosphorylation associated proteins had an increased expression in PAH-EC compared to CTEPH-EC and HPAEC. PAH-EC, CTEPH-EC and HPAEC presented similar HOXD macrovascular gene expression. Metabolic inhibitors showed a dose-dependent reduction in viability in all three groups, predominantly in PAH-EC. A different metabolic profile is present in CTEPH-EC compared to PAH-EC and suggests differences in molecular mechanisms important in the disease pathology and treatment.

Asunto(s)

Células Endoteliales/metabolismo , Hipertensión Arterial Pulmonar/genética , Hipertensión Arterial Pulmonar/metabolismo , Embolia Pulmonar/genética , Embolia Pulmonar/metabolismo , Adulto , Anciano , Células Cultivadas , Enfermedad Crónica , Femenino , Expresión Génica , Glutamato Deshidrogenasa/genética , Glutamato Deshidrogenasa/metabolismo , Glucólisis/genética , Hexoquinasa/genética , Hexoquinasa/metabolismo , Humanos , L-Lactato Deshidrogenasa/genética , L-Lactato Deshidrogenasa/metabolismo , Masculino , Persona de Mediana Edad , Fosforilación Oxidativa , Arteria Pulmonar/citología , Piruvato Deshidrogenasa (Lipoamida)/genética , Piruvato Deshidrogenasa (Lipoamida)/metabolismo

RESUMEN

Platelets play a critical role in arterial thrombosis. Rutaecarpine (RUT) was purified from Tetradium ruticarpum, a well-known Chinese medicine. This study examined the relative activity of RUT with NF-κB inhibitors in human platelets. BAY11-7082 (an inhibitor of IκB kinase [IKK]), Ro106-9920 (an inhibitor of proteasomes), and RUT concentration-dependently (1-6 µM) inhibited platelet aggregation and P-selectin expression. RUT was found to have a similar effect to that of BAY11-7082; however, it exhibits more effectiveness than Ro106-9920. RUT suppresses the NF-κB pathway as it inhibits IKK, IκBα, and p65 phosphorylation and reverses IκBα degradation in activated platelets. This study also investigated the role of p38 and NF-κB in cell signaling events and found that SB203580 (an inhibitor of p38) markedly reduced p38, IKK, and p65 phosphorylation and reversed IκBα degradation as well as p65 activation in a confocal microscope, whereas BAY11-7082 had no effects in p38 phosphorylation. The 2,2-diphenyl-1-picrylhydrazyl (DPPH) assay shows that RUT and BAY11-7082 did not exhibit free radical scavenging activity. In the in vivo study, compared with BAY11-7082, RUT more effectively reduced mortality in adenosine diphosphate (ADP)-induced acute pulmonary thromboembolism without affecting the bleeding time. In conclusion, a distinctive pathway of p38-mediated NF-κB activation may involve RUT-mediated antiplatelet activation, and RUT could act as a strong prophylactic or therapeutic drug for cardiovascular diseases.

Asunto(s)

Fibrinolíticos/farmacología , Alcaloides Indólicos/farmacología , FN-kappa B/metabolismo , Nitrilos/farmacología , Quinazolinas/farmacología , Sulfonas/farmacología , Trombosis/tratamiento farmacológico , Trombosis/metabolismo , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismo , Animales , Modelos Animales de Enfermedad , Fibrinolíticos/uso terapéutico , Depuradores de Radicales Libres/farmacología , Depuradores de Radicales Libres/uso terapéutico , Radicales Libres/antagonistas & inhibidores , Humanos , Quinasa I-kappa B/antagonistas & inhibidores , Imidazoles/farmacología , Imidazoles/uso terapéutico , Alcaloides Indólicos/uso terapéutico , Masculino , Ratones Endogámicos ICR , FN-kappa B/antagonistas & inhibidores , Nitrilos/uso terapéutico , Selectina-P/metabolismo , Activación Plaquetaria/efectos de los fármacos , Inhibidores de Proteínas Quinasas/farmacología , Inhibidores de Proteínas Quinasas/uso terapéutico , Embolia Pulmonar/tratamiento farmacológico , Embolia Pulmonar/metabolismo , Piridinas/farmacología , Piridinas/uso terapéutico , Quinazolinas/uso terapéutico , Sulfonas/uso terapéutico , Factor de Transcripción ReIA/metabolismo

RESUMEN

BACKGROUND: Post-pulmonary embolism (PE) syndrome occurs in up to 50% of PE patients. The pathophysiology of this syndrome is obscure. OBJECTIVE: We investigated whether enhanced oxidative stress and prothrombotic state may be involved in post-PE syndrome. METHODS: We studied 101 normotensive noncancer PE patients (aged 56.5 ± 13.9 years) on admission, after 5-7 days and after a 3-month anticoagulation, mostly with rivaroxaban. A marker of oxidative stress, 8-isoprostane, endogenous thrombin potential, fibrinolysis proteins, clot lysis time (CLT) and fibrin clot permeability (Ks ), along with PE biomarkers, were determined. RESULTS: Patients who developed the post-PE syndrome (n = 31, 30.7%) had at baseline 77.6% higher N-terminal brain natriuretic propeptide and 46.8% higher growth differentiation factor 15, along with 14.1% longer CLT associated with 34.4% higher plasminogen activator inhibitor-1 as compared to subjects without post-PE syndrome (all P < .05). After 5-7 days, only hypofibrinolysis was noted in post-PE syndrome patients. When measured at 3 months, prolonged CLT and reduced Ks were observed in post-PE syndrome patients, accompanied by 23.8% higher growth differentiation factor 15 and 35.8% higher plasminogen activator inhibitor-1 (all P < .05). 8-isoprostane levels ≥108 pg/ml (odds ratio=4.36; 95% confidence interval 1.63-12.27) and growth differentiation factor 15 ≥ 1529 pg/ml (odds ratio=3.89; 95% confidence interval 1.29-12.16) measured at 3 months were associated with higher risk of developing post-PE syndrome. CONCLUSIONS: Enhanced oxidative stress and prothrombotic fibrin clot properties could be involved in the pathogenesis of the post-PE syndrome. Elevated growth differentiation factor 15 assessed at 3 months might be a new biomarker of this syndrome.

Asunto(s)

Dinoprost/análogos & derivados , Factor 15 de Diferenciación de Crecimiento/sangre , Embolia Pulmonar/sangre , Adulto , Anciano , Biomarcadores/sangre , Dinoprost/sangre , Femenino , Humanos , Masculino , Persona de Mediana Edad , Estrés Oxidativo , Embolia Pulmonar/complicaciones , Embolia Pulmonar/metabolismo , Síndrome , Trombosis/complicaciones , Trombosis/metabolismo

RESUMEN

BACKGROUND: The mechanism of chronic thromboembolic pulmonary hypertension (CTEPH) is known to be multifactorial but remains incompletely understood. METHODS: In this study, single-cell RNA sequencing, which facilitates the identification of molecular profiles of samples on an individual cell level, was applied to investigate individual cell types in pulmonary endarterectomized tissues from 5 patients with CTEPH. The order of single-cell types was then traced along the developmental trajectory of CTEPH by trajectory inference analysis, and intercellular communication was characterized by analysis of ligand-receptor pairs between cell types. Finally, comprehensive bioinformatics tools were used to analyze possible functions of branch-specific cell types and the underlying mechanisms. RESULTS: Eleven cell types were identified, with immune-related cell types (T cells, natural killer cells, macrophages, and mast cells) distributed in the left (early) branch of the pseudotime tree, cancer stem cells, and CRISPLD2+ cells as intermediate cell types, and classic disease-related cell types (fibroblasts, smooth muscle cells, myofibroblasts, and endothelial cells) in the right (later) branch. Ligand-receptor interactions revealed close communication between macrophages and disease-related cell types as well as between smooth muscle cells and fibroblasts or endothelial cells. Moreover, the ligands and receptors were significantly enriched in key pathways such as the PI3K/Akt signaling pathway. Furthermore, highly expressed genes specific to the undefined cell type were significantly enriched in important functions associated with regulation of endoplasmic reticulum stress. CONCLUSIONS: This single-cell RNA sequencing analysis revealed the order of single cells along a developmental trajectory in CTEPH as well as close communication between different cell types in CTEPH pathogenesis.

Asunto(s)

Células Endoteliales/metabolismo , Hipertensión Pulmonar/metabolismo , Pulmón/metabolismo , Embolia Pulmonar/metabolismo , Humanos , Macrófagos/metabolismo , Miocitos del Músculo Liso/metabolismo , Arteria Pulmonar/metabolismo , Transducción de Señal/fisiología

RESUMEN

[Figure: see text].

Asunto(s)

COVID-19/metabolismo , Inflamación/metabolismo , Accidente Cerebrovascular Isquémico/metabolismo , Trombofilia/metabolismo , Anciano , Anciano de 80 o más Años , Sedimentación Sanguínea , Proteína C-Reactiva/metabolismo , COVID-19/complicaciones , Análisis por Conglomerados , Femenino , Ferritinas/metabolismo , Productos de Degradación de Fibrina-Fibrinógeno/metabolismo , Fibrinógeno/metabolismo , Mortalidad Hospitalaria , Humanos , Interleucina-6/metabolismo , Accidente Cerebrovascular Isquémico/complicaciones , L-Lactato Deshidrogenasa/metabolismo , Recuento de Leucocitos , Modelos Logísticos , Aprendizaje Automático , Masculino , Persona de Mediana Edad , Infarto del Miocardio/complicaciones , Infarto del Miocardio/metabolismo , Tiempo de Tromboplastina Parcial , Embolia Pulmonar/complicaciones , Embolia Pulmonar/metabolismo , Estudios Retrospectivos , SARS-CoV-2 , Índice de Severidad de la Enfermedad , Trombosis de la Vena/complicaciones , Trombosis de la Vena/metabolismo

RESUMEN

Venous thromboembolism is the third common cardiovascular disease and is composed of two entities, deep vein thrombosis (DVT) and its potential fatal form, pulmonary embolism (PE). While PE is observed in ~ 40% of patients with documented DVT, there is limited biomarkers that can help identifying patients at high PE risk. To fill this need, we implemented a two hidden-layers artificial neural networks (ANN) on 376 antibodies and 19 biological traits measured in the plasma of 1388 DVT patients, with or without PE, of the MARTHA study. We used the LIME algorithm to obtain a linear approximate of the resulting ANN prediction model. As MARTHA patients were typed for genotyping DNA arrays, a genome wide association study (GWAS) was conducted on the LIME estimate. Detected single nucleotide polymorphisms (SNPs) were tested for association with PE risk in MARTHA. Main findings were replicated in the EOVT study composed of 143 PE patients and 196 DVT only patients. The derived ANN model for PE achieved an accuracy of 0.89 and 0.79 in our training and testing sets, respectively. A GWAS on the LIME approximate identified a strong statistical association peak (rs1424597: p = 5.3 × 10-7) at the PLXNA4 locus. Homozygote carriers for the rs1424597-A allele were then more frequently observed in PE than in DVT patients from the MARTHA (2% vs. 0.4%, p = 0.005) and the EOVT (3% vs. 0%, p = 0.013) studies. In a sample of 112 COVID-19 patients known to have endotheliopathy leading to acute lung injury and an increased risk of PE, decreased PLXNA4 levels were associated (p = 0.025) with worsened respiratory function. Using an original integrated proteomics and genetics strategy, we identified PLXNA4 as a new susceptibility gene for PE whose exact role now needs to be further elucidated.

Asunto(s)

Predisposición Genética a la Enfermedad/genética , Redes Neurales de la Computación , Proteómica , Embolia Pulmonar/sangre , Embolia Pulmonar/genética , Receptores de Superficie Celular/sangre , Receptores de Superficie Celular/genética , Adulto , COVID-19/complicaciones , Femenino , Estudio de Asociación del Genoma Completo , Humanos , Masculino , Fenotipo , Polimorfismo de Nucleótido Simple , Embolia Pulmonar/complicaciones , Embolia Pulmonar/metabolismo

RESUMEN

This study was intended to investigate the effect of Curcumin on acute pulmonary embolism (APE) via microRNA-21 (miR-21)/PTEN/NF-κB axis. APE model was induced on rats and administrated with Curcumin. Western blot analysis and RT-qPCR manifested the downregulation of Sp1, miR-21 and NF-κB, but the upregulation of PTEN in Curcumin-treated APE rats. Blood gas analysis, ELISA, and weighing of wet weight/dry weight (W/D) ratio indicated that Curcumin diminished mPAP and RVSP levels, W/D ratio, thrombus volume, and inflammatory factors in the lungs of APE rats. Further mechanical analysis was conducted by dual-luciferase reporter assays and ChIP assay, which showed that Sp1 increased miR-21 expression by binding to the miR-21 promoter, and that PTEN was targeted by miR-21. The APE rats were injected with adenovirus to evaluate the effect of Sp1, miR-21, or PTEN on lung injury and inflammation. It was observed that downregulation of miR-21 or Sp1, or upregulation of PTEN diminished mPAP and RVSP levels, W/D ratio, thrombus volume, and inflammatory factors in the lungs of APE rats. In summary, Curcumin decreased miR-21 expression by downregulating Sp1 to upregulate PTEN and to impair the NF-κB signaling pathway, thus suppressing lung injury and inflammation in APE rats.

Asunto(s)

Lesión Pulmonar Aguda , Curcumina/farmacología , MicroARNs/metabolismo , FN-kappa B/metabolismo , Fosfohidrolasa PTEN/metabolismo , Embolia Pulmonar , Transducción de Señal/efectos de los fármacos , Lesión Pulmonar Aguda/metabolismo , Lesión Pulmonar Aguda/prevención & control , Animales , Inflamación/metabolismo , Inflamación/prevención & control , Embolia Pulmonar/metabolismo , Embolia Pulmonar/prevención & control , Ratas , Ratas Sprague-Dawley

RESUMEN

Chronic thromboembolic pulmonary hypertension (CTEPH) is a vascular disease characterized by the presence of organized thromboembolic material in pulmonary arteries leading to increased vascular resistance, heart failure and death. Dysfunction of endothelial cells is involved in CTEPH. The present study describes for the first time the molecular processes underlying endothelial dysfunction in the development of the CTEPH. The advanced analytical approach and the protein network analyses of patient derived CTEPH endothelial cells allowed the quantitation of 3258 proteins. The 673 differentially regulated proteins were associated with functional and disease protein network modules. The protein network analyses resulted in the characterization of dysregulated pathways associated with endothelial dysfunction, such as mitochondrial dysfunction, oxidative phosphorylation, sirtuin signaling, inflammatory response, oxidative stress and fatty acid metabolism related pathways. In addition, the quantification of advanced oxidation protein products, total protein carbonyl content, and intracellular reactive oxygen species resulted increased attesting the dysregulation of oxidative stress response. In conclusion this is the first quantitative study to highlight the involvement of endothelial dysfunction in CTEPH using patient samples and by network medicine approach.

Asunto(s)

Células Endoteliales/metabolismo , Hipertensión Pulmonar/metabolismo , Carbonilación Proteica , Mapas de Interacción de Proteínas , Arteria Pulmonar/metabolismo , Embolia Pulmonar/metabolismo , Tromboembolia/metabolismo , Células Endoteliales/patología , Humanos , Hipertensión Pulmonar/patología , Arteria Pulmonar/patología , Embolia Pulmonar/patología , Tromboembolia/patología