11,972 research outputs found

    Netrin-1 isoforms and macrophage phenotype: role in the pathophysiology of atherosclerosis

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    Atherosclerosis is a chronic inflammatory disease and is the main cause of ischaemic heart disease, stroke, and other cardiovascular diseases. The development of atherosclerotic plaques incidence has been increasing significantly during the last few decades and is due to different factors, such as the increased consumption of high fat food, smoking and increased incidence of diabetes mellitus. This is currently a major burden on health systems, which require new therapeutic targets to address this challenge. The involvement of netrin-1 in inflammation and atherosclerosis has been studied during the last two decades. Different groups have established that netrin-1 plays a prominent role in these contexts and has different effects depending on where it is produced, and which cells are targeted by it. While endothelial-derived netrin-1 secreted into the circulation gives rise to a protective effect against atherosclerosis, macrophage-derived netrin-1 within the plaque has a pro-atherogenic effect, promoting the trapping and survivability of foam cells. More recently, a truncated isoform of netrin-1 was found in the nuclei of different types of cancer cells. This form of netrin-1 has also been studied in endothelial cells, but so far, no relationship between the expression of truncated netrin-1 and macrophages has been established. The link between cytokine stimulation, macrophage phenotype and netrin-1 isoforms has until now been unclear. The work described in this thesis looked at the expression and function of netrin-1 on monocytes and different macrophage phenotypes. Gene expression analysis revealed that macrophages express both full-length and truncated netrin-1. Classical activated is the macrophage phenotype that presents higher expression of netrin-1, and the expression of both isoforms is at least partially dependent on NF-κB activation pathway. Netrin-1 inhibits monocyte migration by inhibiting chemokines’ chemoattractant effect. CCL2 and netrin-1 individually showed an attractant effect towards THP-1-derived macrophages, but the signals were inhibited when the two were combined. The apoptotic agent selected to study netrin-1 anti-apoptotic effect was not UNC5b-dependent and, therefore, limited our ability to acquire relevant data regarding this process. Administration of exogenous netrin-1 to mice, increasing its systemic levels, showed an acute and chronic protective effect against inflammation. Mice treated with netrin-1 showed less macrophages within the tissue after the induction of local inflammation. Furthermore, increasing the netrin-1 systemic levels of mice prevented the enlargement of the aortic sinus and development of plaque after feeding them 60% HFD over a 6-week period. This thesis provides relevant insights into the role of macrophage phenotypes in the expression of netrin-1 isoforms within atherosclerotic plaques, and how netrin-1 affects these immune cells. The findings highlight the significance of netrin-1 as a potential therapeutic target for treating not only cardiovascular disease but also other inflammatory conditions. Therefore, this research sheds light on the promising future of netrin-1 as a treatment option for a range of diseases

    Malignant pleural effusion: current understanding and therapeutic approach

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    Malignant pleural effusion (MPE) is a common complication of thoracic and extrathoracic malignancies and is associated with high mortality and elevated costs to healthcare systems. Over the last decades the understanding of pathophysiology mechanisms, diagnostic techniques and optimal treatment intervention in MPE have been greatly advanced by recent high-quality research, leading to an ever less invasive diagnostic approach and more personalized management. Despite a number of management options, including talc pleurodesis, indwelling pleural catheters and combinations of the two, treatment for MPE remains symptom directed and centered around drainage strategy. In the next future, because of a better understanding of underlying tumor biology together with more sensitive molecular diagnostic techniques, it is likely that combined diagnostic and therapeutic procedures allowing near total outpatient management of MPE will become popular. This article provides a review of the current advances, new discoveries and future directions in the pathophysiology, diagnosis and management of MPE

    Off-pump minimally invasive coronary artery bypass grafting in patients with left ventricular dysfunction: the lampang experience

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    IntroductionThe minimally invasive cardiac surgery off-pump coronary artery bypass (MICSOPCAB) is technically difficult; therefore, previous studies have indicated that MICSOPCAB should be contraindicated in patients with impaired left ventricular (LV) function. In this study, we investigated the feasibility of MICSOPCAB in patients with impaired LV function.MethodsThe 226 patients underwent MICSOPCAB between August 2017 and September 2022. Our study defined impaired LV function as ejection fraction (EF) in echocardiography 40% or less. The patients were divided into Low EF group (n = 39) and Normal EF group (n = 187).ResultsThe Low EF group was in a more critical preoperative condition than Normal EF group (41.0% in the Low EF group vs. 14.4% in the Normal EF group; p < 0.001). For preoperative transthoracic echocardiography, LV end-diastolic diameter (5.5 ± 0.9 cm in the Low EF group vs. 5.0 ± 0.8 cm in the Normal EF group; p < 0.001) and LV end-systolic diameter (4.4 ± 1.0 cm in the Low EF group vs. 3.4 ± 1.0 cm in the Normal EF group; p < 0.001) were significantly larger in the Low EF group. No differences were found in the operative time (180 [160–240] min in the Low EF group vs. 205 [165–253] min in the Normal EF group; p = 0.231) and the median number of distal anastomoses (2 [1–2] in the Low EF group vs. 2 [1–3] in the Normal EF group; p = 0.073). Intensive care unit stay was longer in the Low EF group than in the Normal EF group (2 [1–2] in the Low EF group vs. 1 [1–2] in the Normal EF group; p = 0.010). Perioperative transfusion was more common in the Low EF group than in the Normal EF group (69.7% vs. 49.2%; p = 0.023). There were no differences in major complications, hospital stay, and 30-day mortality. The Kaplan–Meier curve showed no significant difference in postoperative major adverse cardiac or cerebrovascular events rates between the two groups (p = 0.185)ConclusionIn this study, MICSOPCAB can be performed in patients with low EF having short- and mid-term outcomes similar to patients with normal EF. Therefore, low EF should not be contraindicated in MICSOPCAB

    Role of calcium-activated potassium channels in the regulation of basal and agonist-elevated tones in isolated conduit arteries

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    Functional role of calcium-activated potassium (KCA) channels on the basal and agonist-elevated arterial tones was investigated in isolated rabbit aorta, porcine and canine coronary arteries as well as in human internal mammary artery. The vascular tones enhanced by contractile agents were increased further by preincubation of these conduit blood vessels with selective (charybdotoxin or iberiotoxin) or non-selective (tetraethylammonium) inhibitors of KCA channels. The basal tone (without an agonist) was increased only in the canine coronary artery. The results indicate a feed-back regulatory role of KCA channels counteracting the vasospasm of conduit arteries

    Feasibility of Dose Reduction to the Left Anterior Descending Coronary Artery without Compromising Target Volume Coverage Using Tomotherapy Techniques

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    Background: Radiotherapy is associated with a high risk of heart disease in patients with left-sided breast cancer. Previously, the entire heart was considered an organ at risk (OAR) during planning. Studies have shown that the effect of radiation therapy depends on the dose to specific heart substructures. However, the tolerance dose of the left anterior descending coronary artery (LAD), an important cardiac substructure, is yet to be determined. This study aims to verify the feasibility of reducing the LAD dose with appropriate dose-volume constraints for patients undergoing left whole-breast radiotherapy, without compromising the target dose coverage, using tomotherapy techniques.Method: This retrospective study generated tomohelical and tomodirect plans initially without considering the LAD as OAR in the treatment planning. To reduce the LAD dose, plans were regenerated by including the LAD as an OAR with appropriate dose constraints. The dose-volume histogram parameters of these plans were compared with those of the initial plans of the respective types.Results: Tomohelical plans showed a 4.4% reduction in maximum dose and a 3.8% reduction in V15 for LAD, while tomodirect plans registered a 3% reduction in V15, with the conformity index remaining constant. Based on the LAD dosimetric results, considering the LAD as an OAR is associated with lower LAD doses without compromising the target volume coverage.Conclusion: It is feasible to reduce the LAD dose without compromising target volume coverage or affecting other OAR doses in patients with left breast cancer, using tomotherapy techniques

    Fabrication of tubular constructs using hybrid extrusion printing and electrospinning for vascular tissue engineering applications

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    Cardiovascular disease is the leading cause of mortality in the world, with the number of deaths rising every year. Coronary artery disease, peripheral artery disease, aneurysm, and strokes are the most prominent forms of vascular diseases. Bypass surgery is one of the techniques performed to treat these vascular diseases, which involves the re-routing of blood flow around the blocked arterial site using a vascular graft. Autologous grafts, such as saphenous vein and internal mammary artery, remain a gold standard for bypass grafting procedures. However, the removal of autologous grafts often results in the donor site morbidity. In some cases, the grafts are unavailable due to previous harvest or poor quality. Synthetic grafts represent an alternate option, however, they usually fail when used as small diameter bypass grafts. The mismatch between the mechanical properties of synthetic grafts and native arteries is the major cause of failure of these synthetic grafts made from PET and ePTFE. Therefore, there is an urgent and high demand to fabricate a vascular conduit having biomechanical behaviour similar to that of the native arteries. In this thesis, a new hybrid extrusion printing and electrospinning technique is presented to fabricate vascular grafts. A commercially available Creality3D Ender 3D printer was modified into a hybrid setup having a vertical mandrel, an extrusion printing head, and electrospinning heads. The setup is capable of fabricating layered structure using hydrogel and electrospun nanofibres. The gelatin methacryloyl (gelMA) hydrogel was selected for printing tubular constructs as it is widely used in the literature to encapsulate cells for bioprinting. Polycaprolactone (PCL) and Poly (L-lactide-co-ε- caprolactone) (PLCL) polymers were used to produce electrospun nanofibres for the reinforcement of hydrogel conduits. The setup was successfully tested to print grafts around the rotating mandrel. Bioprinting is an attractive technique to print tissues using hydrogels. However, printing of long tubular constructs from hydrogels remains a challenge. Additive-lathe printing method offers a solution to print grafts with high aspect ratios but printing hydrogels around the horizontal rods often leads to sagging, which results in the non-uniform wall thickness and subsequently variable mechanical properties. A new approach of printing around the vertical mandrel was used to fabricate gelMA grafts having uniform wall thickness. The printing parameters were selected to achieve the overlapping of two consecutive printed filaments, which resulted in the better bonding of filaments and good quality tubular constructs were fabricated. The tensile testing results revealed that the anisotropic properties of printed gelMA grafts were similar to those observed in the natural blood vessels. Moreover, no leakage was detected in the printed gelMA tubular constructs during the burst pressure measurement. Thus, a vertical additive-lathe printing method offers an attractive technique to print long vascular grafts using hydrogels. The burst pressure of the gelMA grafts printed on vertical additive-lathe printing setup was found to be lower than the normal haemodynamic blood pressures. Therefore, a reinforcement is required. A layer of electrospun nanofibres was collected over the printed hydrogel constructs to improve the mechanical performance. Various blends of Polycaprolactone (PCL) and poly(L-lactide-co-ε-caprolactone) (PLCL) polymer solutions were used to produce electrospun nanofibres. The printed bi-layered vascular grafts showed mechanical properties close to that of the native arteries. The tensile strength of the 12% gelMA constructs reinforced by 100/0 PCL/PLCL blend fibres was found to be greater than 3 MPa, which falls in the range of tensile strength value of native arteries (0.5 – 3 MPa for coronary arteries and 1.5 – 4 MPa for radial and mammary arteries). The fabricated vascular grafts showed a burst pressure of more than 2000 mmHg, which is within the range of burst pressure values for the human saphenous vein (1250 – 2476 mmHg) and radial artery (2001 – 2476 mmHg). Moreover, the compliance values of gelMA constructs reinforced by 100/0 PCL/PLCL (6.85 ± 1.01 %/100mmHg) and 75/25 PCL/PLCL (17.13 ± 7.35 %/100mmHg) were found to be similar to the muscular arteries (6.03 ± 3.39 %/100mmHg) and elastic arteries (16.21 ± 3.81 %/100mmHg), respectively. The cytocompatibility assessment showed that gelMA presented a bioactive surface for the endothelial cells to survive and grow. The newly developed hybrid setup has been utilized to fabricate fibre-reinforced vascular grafts. The mechanical performance of these constructs was found to be similar to that of native arteries. Also, PCL/PLCL fibres showed good cellular metabolic activity and gelMA surfaces were biocompatible for the HUVECs to grow. Thus, the presented fabrication method has a great potential to fabricate vascular grafts having biomechanical properties similar to that of natural blood vessels

    Enhanced hemocompatibility and rapid magnetic anastomosis of electrospun small-diameter artificial vascular grafts

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    Background: Small-diameter (<6 mm) artificial vascular grafts (AVGs) are urgently required in vessel reconstructive surgery but constrained by suboptimal hemocompatibility and the complexity of anastomotic procedures. This study introduces coaxial electrospinning and magnetic anastomosis techniques to improve graft performance.Methods: Bilayer poly(lactide-co-caprolactone) (PLCL) grafts were fabricated by coaxial electrospinning to encapsulate heparin in the inner layer for anticoagulation. Magnetic rings were embedded at both ends of the nanofiber conduit to construct a magnetic anastomosis small-diameter AVG. Material properties were characterized by micromorphology, fourier transform infrared (FTIR) spectra, mechanical tests, in vitro heparin release and hemocompatibility. In vivo performance was evaluated in a rabbit model of inferior vena cava replacement.Results: Coaxial electrospinning produced PLCL/heparin grafts with sustained heparin release, lower platelet adhesion, prolonged clotting times, higher Young’s modulus and tensile strength versus PLCL grafts. Magnetic anastomosis was significantly faster than suturing (3.65 ± 0.83 vs. 20.32 ± 3.45 min, p < 0.001) and with higher success rate (100% vs. 80%). Furthermore, magnetic AVG had higher short-term patency (2 days: 100% vs. 60%; 7 days: 40% vs. 0%) but similar long-term occlusion as sutured grafts.Conclusion: Coaxial electrospinning improved hemocompatibility and magnetic anastomosis enhanced implantability of small-diameter AVG. Short-term patency was excellent, but further optimization of anticoagulation is needed for long-term patency. This combinatorial approach holds promise for vascular graft engineering

    Effects of municipal smoke-free ordinances on secondhand smoke exposure in the Republic of Korea

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    ObjectiveTo reduce premature deaths due to secondhand smoke (SHS) exposure among non-smokers, the Republic of Korea (ROK) adopted changes to the National Health Promotion Act, which allowed local governments to enact municipal ordinances to strengthen their authority to designate smoke-free areas and levy penalty fines. In this study, we examined national trends in SHS exposure after the introduction of these municipal ordinances at the city level in 2010.MethodsWe used interrupted time series analysis to assess whether the trends of SHS exposure in the workplace and at home, and the primary cigarette smoking rate changed following the policy adjustment in the national legislation in ROK. Population-standardized data for selected variables were retrieved from a nationally representative survey dataset and used to study the policy action’s effectiveness.ResultsFollowing the change in the legislation, SHS exposure in the workplace reversed course from an increasing (18% per year) trend prior to the introduction of these smoke-free ordinances to a decreasing (−10% per year) trend after adoption and enforcement of these laws (β2 = 0.18, p-value = 0.07; β3 = −0.10, p-value = 0.02). SHS exposure at home (β2 = 0.10, p-value = 0.09; β3 = −0.03, p-value = 0.14) and the primary cigarette smoking rate (β2 = 0.03, p-value = 0.10; β3 = 0.008, p-value = 0.15) showed no significant changes in the sampled period. Although analyses stratified by sex showed that the allowance of municipal ordinances resulted in reduced SHS exposure in the workplace for both males and females, they did not affect the primary cigarette smoking rate as much, especially among females.ConclusionStrengthening the role of local governments by giving them the authority to enact and enforce penalties on SHS exposure violation helped ROK to reduce SHS exposure in the workplace. However, smoking behaviors and related activities seemed to shift to less restrictive areas such as on the streets and in apartment hallways, negating some of the effects due to these ordinances. Future studies should investigate how smoke-free policies beyond public places can further reduce the SHS exposure in ROK

    Stroke dependent mechanisms of endothelial autoimmune PAR2 activation

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    Protease-activated receptor 2 (PAR2) is broadly expressed on endothelial cells contributing to inflammatory processes and angiogenesis. Autoantibodies against PAR2 can regulate these functions in health and disease. Recently, a collaboration group discovered in a prospective observational cohort study that stroke patients with a higher concentration (highest quartile Q4) of PAR2-IgG presented a poor clinical outcome compared to others (lower quartiles Q1-3). However, the biological functions and molecular mechanism remain unclear. Therefore this study aims to explore the possible function of PAR2-IgG on endothelial-based angiogenesis which might have an essential impact on clinical outcome after stroke. Methods: IgGs were isolated from the sera of stroke patients (Q1-3 and Q4), then Matrigel angiogenesis assay was used to study vascular tube formation by IgGs and the natural PAR2 agonist trypsin in vascular endothelial cells (HMEC-1 and hCMEC/D3, a blood-brain barrier human endothelial cell line). Additionally, qRT-PCR, western blot, and ELISA were used to investigate the angiogenic signaling pathways. The activation of G-protein subunits was examined by the Luciferase reporter assay, based on mutations of wild-type PAR2 and extracellular loops (ECLs). The receptor internalization was studied by Nano-Glo ®HiBiT detection assay. The main inductor of angiogenesis VEGF was examined in detail by promoter assay with regard to the activation of the promoter and potential binding fragments of transcription factors. Results: The PAR2 agonist trypsin was involved in the regulation of mRNA synthesis and protein release of VEGF, IL-6, IL-8, CXCL1 and ANG2 through the ERK1/2 signaling pathway, which promoted vascular tube formation of HMEC-1. Q1-3 PAR2-IgG enhanced angiogenesis via PAR2-ERK1/2-induced angiogenic cytokines and receptor internalization when compared to Q4 PAR2-IgG. Q1-3 PAR2-IgG 10 triggered Gq/11 activation -, but not G12/13 -, and the mutation of ECL1-3 did not alter the activation. As for Q4, the ECL3 mutation increased the Gq/11 activation and ERK1/2 activation. Different truncations of the VEGF promoter assay confirmed the binding sites of transcription factors located in the VEGF promoter sequence -1339 to -839 bp, and the inhibition of AP-1 completely blocked the promoter activation. To summarize, in contrast to the Q4 group, Q1-3 PAR2-IgG induced greater receptor internalization, Gq/11 activation, ERK1/2 activation, and angiogenic gene production, which resulted in increased formation of vascular tubes. Conclusion: The present work elucidated new PAR2 signaling pathways that are induced by PAR2-IgG in human endothelial cells and identified contrary functional consequences on the angiogenesis of PAR2-IgGs from two different stroke patient cohorts classified by different PAR2-IgG titer ranges. Novel therapeutics modulating PAR2-IgGs in stroke patients might thereby already limit stroke progression at an early stage and lead to improved clinical outcome.Hintergrund: Protease-aktivierter Rezeptor 2 (PAR2) ist auf Endothelzellen hoch exprimiert und reguliert inflammatorische Prozesse und Angiogenese. Autoantikörper gegen PAR2 können diese Funktionen physiologisch und pathophysiologisch beeinflussen. Vor kurzem entdeckten Kollaborationspartner in einer prospektiven Kohortenstudie, dass Schlaganfallpatienten mit hohen PAR2-Spiegeln (höchstes Quartil Q4) einen schlechteren klinischen Outcome hatten als Patienten mit niedrigeren Spiegeln (Quartile Q1-3). Die molekularen Mechanismen dafür waren allerdings ungeklärt. Es war daher das Ziel dieser Studie, eine mögliche Bedeutung dieser PAR2-IgGs auf endotheliale Angiogenese zu untersuchen, die sich positiv auf den klinischen Outcome auswirken könnten. Methoden: IgGs wurden von Schlaganfallpatienten (Q1-3 und Q4) isoliert und dann der Matrigel Angiogenese Assay benutzt, um Gefäßbildungen von Endothelzellen (HMEC-1 und hCMEC/D3, einer humanen Bluthirnschranken-Endothelzelllinie) durch die IgGs und den natürlichen Agonisten von PAR2, Trypsin, zu untersuchen. Weiterhin dienten qRT-PCR, Westernblots und ELISA zur Erforschung angiogenetischer Signalwege. Luziferase-Reporter-Assays wurden durchgeführt, um die Aktivierung von G-Protein-Untereinheiten mithilfe von Mutationen von Wildtyp-PAR2 und der extrazellulären Schleifen (ECLs) zu untersuchen. Die Internalisierung des Rezeptors wurde visualisiert mit dem Nano-Glo ®HiBiT Detektionsassay. Der Hauptaktivator der Angiogenese, VEGF, wurde detailliert untersucht mithilfe von Promotoraktivierungsstudien und Bindungsfragmente. Ergebnisse: Der PAR2 Agonist Trypsin beeinflusste die mRNA Synthese und Proteinfreisetzung von VEGF, IL-6, IL-8, CXCL1 und ANG2 über den ERK1/2 Signalweg und förderte die Ausbildung vaskulärer Gefäßschläuche in HMEC-1. Im Vergleich zu Q4 PAR2-IgG erhöhte Q1-3 PAR2-IgG Angiogenese durch Freisetzung von 13 angiogentischer Zytokine über PAR2-ERK1/2-Aktivierung und Rezeptorinternalisierung. Q1-3 PAR2-IgG triggerte die Aktivierung von Gq/11, aber nicht von G12/13, und Mutationen von ECL1-3 hatten keinen Einfluss auf die Aktivierung. VEGF Promotorassays bestätigten die Bindungsstelle des Transkriptionsfaktors AP-1 an der VEGF-Promotorsequenz -1339 bis -839 bp. Zusammenfassend induzierte Q1-3 PAR2-IgG im Gegensatz zu Q4 PAR2-IgG eine verstärkte Rezeptorinternalisierung, Gq/11 – und ERK1/2-Aktivierung und Induktion angiogenetischer Gene, was in einer verstärkten Netzwerkbildung resultierte. Schlussfolgerung: Die hier dargestellte Arbeit beschreibt neue Signalwege von PAR2 in humanen Endothelzellen, die durch PAR2-IgG aktiviert werden und identifizierte gegensätzliche Auswirkungen auf Angiogenese durch PAR2-IgG von zwei unterschiedlichen Schlaganfallkohorten mit besonders hohen (Q4) oder niedrig-mäßigen (Q1-3) PAR2-IgG-Spiegeln. Die Modulation von PAR2-IgGs könnte eine neue Therapieform bei Schlaganfall darstellen, um die Progression im Verlauf zu verhindern und zu verbessertem klinischem Outcome zu führen
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