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5‐羟色胺（5-HT）是一种强有力的血管活性神经递质，被广泛的应用在调节血管张力。当5‐HT 被释放后，会被单胺氧化酶（MAOs）催化的酶促反应代谢，从而产生不同的代谢产物，比如5‐HIAA，5‐HTOL 和过氧化氢（H₂O₂）。然而，5‐HT对于内皮细胞活性氧物种（ROS）的产生作用以及5‐HT 转运体，5‐HT 受体，MAOs和ROS 的产生伴随着细胞内钙变化是否参与了其中的信号传导尚未被阐明。所以，这个研究最初的目的是考查外源性加入的5‐HT 对于脐静脉内皮细胞中ROS产生的影响以及其潜在的生物机理。 / 数据清楚的显示在没有L‐NAME（一种抑制一氧化氮（NO）产生的抑制剂）预处理的情况下，5‐HT 并不能在脐静脉内皮细胞内产生显著性的ROS。然而，在L‐NAME 预处理的情况下，NO 的产生被完全抑制，我们观察到明显的显著性的线粒体内的ROS 产生。5‐HT 产生的线粒体ROS 可以被clorgyline（一种MAO‐A 抑制剂），indatraline（一种5‐HT 转运体阻断剂），LY272015（一种5‐HT‐2B 受体拮抗剂），ketanserin（一种5‐HT2A 受体拮抗剂），XeC（一种IP3 受体拮抗剂），Gd³⁺（一种非选择性TRP 通道阻断剂），BAPTA（一种强效钙离子螯合剂），PEG‐Catalase，U73122（一种选择性PLC 抑制剂）以及没有钙离子的培养基所阻止。同时，5‐HT介导的胞内钙离子变化被XeC, Gd³⁺, BAPTA, U73122, ketanserin, LY272015 以及没有钙离子的培养基所阻止。另外，MAO‐A 基因敲除抑制了5‐HT 导致的线粒体ROS的产生却对5‐HT 介导的胞内钙离子变化没有影响。基于以上所述的结果，我们可以得出结论，通过5‐HT 转运体，5‐HT 被摄取入细胞内，然后通过MAO‐A 介导的酶促代谢反应，产生钙离子依赖性的线粒体内ROS 的产生，这一结论对于解释血小板聚集而引起的内皮细胞功能性障碍起到非常重要的作用。 / 根据前人所述，内皮细胞内产生的ROS 对于内皮细胞通透性变化有着重要的作用，但是5‐HT 诱导的脐静脉内皮细胞ROS 的增加是否会对内皮通透性有所影响并没有被说明。在这项研究中，我们设计了实验旨在测试平面细胞表面积（ PCSA ）， 跨内皮电阻（ TER ）， 细胞高度， 肌球蛋白轻链磷酸化（MLCphosphorylation）和肌动蛋白细胞骨架（F‐actin cytoskeleton）水平的变化。此外，b‐catenin 在ROS 引起的F‐actin cytoskeleton 重组中的作用也在我们的讨论范围之内。 / 数据表明，在L‐NAME 预处理的情况下，5-HT 降低了脐静脉内皮的PCSA，TER 以及细胞高度，却增加了MLCP 和与b‐catenin 表达负相关的F‐actincytoskeleton 的水平。这些作用明显被PEG‐Catalase 预处理和MAO‐A 基因敲除减弱，证明了5‐HT 通过MAO‐A 介导产生的H₂O₂ 可以增加内皮细胞的通透性。 / 据文献报道，不论内源性还是外源性的低浓度的H₂O₂ 都可以激活导致血管生成的信号通路。文献进一步表明5‐HT 可以通过特定的5‐HT 受体亚型促进各种类型的内皮细胞的血管生成。然而，5‐HT 诱导的H₂O₂ 对于脐静脉内皮的血管生成作用并没有被报道。我们通过最初的实验先验证5‐HT 对于内皮细胞增殖和迁移的影响，然后我们才去验证H₂O₂ 在其中的作用及其潜在的机理。 / 实验结果表明，在L‐NAME 预处理的情况下，不论是急性（30 分钟）还是慢性（24 小时）的5‐HT 的处理都可以导致脐静脉内皮细胞的迁移，而这个作用会被5‐HT‐2 受体拮抗剂ketanserin，LY272015，ROS 清除剂PEG‐Catalase 以及PI3K的抑制剂wortmannin 所抑制。同时，在L‐NAME 预处理下，5‐HT 增加了cortactin,p‐Akt 和 p‐eNOS 的蛋白表达量而并没有影响Akt, eNOS 和p‐cortactin 的蛋白表达量。而5‐HT 增加的p‐Akt 和p‐eNOS 的蛋白表达被wortmannin 和PEG‐Catalase所抑制。不论是在Cyuant 细胞增殖检测还是在BrdU 细胞增殖检测中，5‐HT 诱导了一种非显著性的DNA 合成的增加，并且再BrdU 细胞增殖检测中，增加了的DNA 合成被PEG‐Catalase 显著性降低。总结以上实验结果，我们可以得出结论，通过一种5‐HT‐2 受体介导的PI3K 依赖性通路，而不是cortactin 磷酸化依赖性的信号通，路5‐HT 可以引导内皮细胞迁移。 / 除此之外，ROS 也被印证可以加剧内皮细胞的炎症反应和加速内皮细胞的老化。因此，我们也观察了5‐HT 对于粘附蛋白比如ICAM‐1 和VCAM‐1 以及抗老化因子SIRT‐1 的表达是否有影响。数据表明，在L‐NAME 预处理的情况下，30 分钟的5‐HT 处理显著的增加了ICAM‐1，SIRT‐1 而不是VCAM‐1 的表达。同时，这些作用均可以被PEG‐Catalase 所抑制表明了5‐HT 通过诱导H₂O₂ 的产生来形式其促进炎症反应和抗衰老的作用。 / 最后，总结以上，通过抑制NO 的产生，5‐HT 可以通过MAO‐A 介导的酶促代谢反应在人体脐静脉内皮细胞线粒体诱导ROS 的产生。同时，5‐HT 诱导的H₂O₂参与了改变内皮细胞通透性，促进血管生成（内皮迁移）及炎症反应的过程。 / 5-Hydroxytryptamine (5-HT), a potent vasoactive neurotransmitter, is involved in the regulation of vascular tone. After its release, 5-HT is terminated at the nerve terminals via enzymatic metabolism catalyzed by monoamine oxidases (MAOs), resulting in the generation of different metabolites (e.g. 5-HIAA, 5-HTOL and H₂O₂). Our lab demonstrates for the first time that 5-HT-induced ROS production indeed occurs and therefore, the aim of this study is to investigate exogenously added 5-HT on ROS generation in human umbilical vein endothelial cells (HUVECs), in order to understand the mechanisms involved in 5-HT-induced ROS production. / Our results clearly demonstrated that in the absence of L-NAME(a NO production inhibitor), there wasno apparent ROS production induced by 5-HT. However, after the inhibition of NO synthesis by L-NAME, 5-HT caused a significant increase in mitochondrial H₂O₂ production. The 5-HT-induced mitochondrial H₂O₂ generation was sensitive to clorgyline (a MAO-A inhibitor), indatraline (a 5-HT transporter blocker), LY272015 (a 5-HT2B antagonist) and ketanserin (a 5-HT2A antagonist), Xextospongin C（XeC，a IP3 receptor antagonist), Gd³⁺ (a non-selective TRP channel blocker), BAPTA (a potent Ca²⁺ ions chelator), PEG-Catalase, U73122 (a selective PLC inhibitor), and in [Ca²⁺]o-free medium. Concurrently, 5-HT-mediated [Ca²⁺]i changes were sensitive to XeC, Gd³⁺, BAPTA, U73122, ketanserin, LY272015, and in [Ca²⁺]o-free conditions. In addition, gene knockdown of MAO-A suppressed 5-HT-elicited H₂O₂ production with no effects on [Ca²⁺]i changes. Based on all the results above, we can conclude that 5-HT caused a Ca²⁺-dependent mitochondrial H₂O₂ generation via MAO-A-mediated metabolism with the pre-requisite uptake of 5-HT into HUVECs through 5-HT transporter. / ROS derived from endothelial cells have been implicated in changes in endothelial permeability, but whether 5-HT-induced H₂O₂ generation could alter endothelial cells permeability has as yet not been demonstrated. Here, we measured the planar cell surface area (PCSA), transendothelial electrical resistance (TER), cell height, myosin light chain phosphorylation and F-actin cytoskeleton level in response to 5-HT challenge to investigate the change of endothelial permeability. Moreover, the participation of β-catenin in regulation of F-actin cytoskeleton remodeling in ROS-modulated alteration in endothelial permeability was also investigated. Results indicated that in the presence of L-NAME, 5-HT reduced the PCSA, TER and cell height in HUVECs. In contrast, 5-HT (with L-NAME) increased myosin light chain phosphorylation (MLCP) expression and F-actin cytoskeleton level, which are negatively associated with β-catenin expression. All of these effects were ameliorated by pre-treatment of PEG-Catalase or gene knockdown of MAO-A, implying 5-HT can consistently elicit the increase in endothelial permeability via MAO-A mediated H₂O₂ generation. / Low dose of ROS from exogenous or endogenous source can activate signaling pathway that lead to angiogenesis.5-HT can promote endothelial angiogenesis through specific 5-HT receptor subtype in various endothelial cell types, but the concomitant ROS generation had not previously been indicated to play a role in the process. In this study, we seek to test out the effects of 5-HT on endothelial cells migration, and should there be a functional role for ROS in the process. / Our results revealed that in the presence of L-NAME, both acute (30 min) and chronic (24 hr) treatment of 5-HT caused HUVECs migration, the effects of which were reversed by pre-incubation of 5-HT-2 receptor antagonists, ketanserin, LY272015, ROS scavenger PEG-Catalase or selective PI3K inhibitor wortmannin. With L-NAME, 5-HT consistently increased cortactin, p-Akt and p-eNOS expression without affecting total Akt, eNOS and p-cortactin protein expression whereas the increased p-Akt and p-eNOS expression are suppressed by pre-treatment of wortmanin or PEG-Catalase. Both in Cyuant cell proliferation assay and BrdU assay, 5-HT caused a trend but non-significant increase in DNA synthesis whereas the pre-treatment of PEG-Catalase significantly suppressed cell proliferation in the BrdU assay. Based on these results, we can conclude that 5-HT elicits endothelial migration via 5-HT-2 receptor-mediated H₂O₂ generation in a PI3K-dependent pathway. Under this circumstance, cortactin phosphorylation-dependent pathway was excluded. / Besides, ROS is notorious for effects like aggravation of inflammation and acceleration aging processes. The investigation extends to looking at alterations ofexpression of adhesion protein including ICAM-1 and VCAM-1 in the inflammatory response pathway and also to looking at the major aging parameter SIRT-1 in the presence of 5-HT in endothelium. Our data showed that in the presence of L-NAME, 30 min treatment of 5-HT significantly increased ICAM-1 and SIRT-1 expression without altering VCAM-1 expression and the up-regulation of ICAM-1 and SIRT-1 expression was prevented by PEG-Catalase. / In conclusion, with the eradication of the influence of NO, 5-HT induced mitochondrial H₂O₂ production via MAO-A-mediated metabolism in HUVECs. At the same time, 5-HT-induced H₂O₂ generation was involved in increasing endothelial permeability, inflammation and angiogenesis (cell migration). / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Zhang, Qian. / Thesis (Ph.D.) Chinese University of Hong Kong, 2013. / Includes bibliographical references (leaves 326-450). / Abstracts also in Chinese. / Zhang, Qian.

Cell-derived microvesicles (MVs) are naturally released into the human circulation and an increase in the concentration of certain MV populations have been observed after exercise. However, the MV appearance dynamics, the exercise-related stimuli that induce their formation and physiological relevance are poorly understood. Hence, the overall objectives of this thesis were to: 1) characterise the circulating platelet (PMV) and endothelial-derived MVs (EMVs) responses during exercise and recovery, as well as their arteriovenous dynamics, 2) investigate the potential role of haemodynamic forces on MVs formation in vivo by vascular shear stress manipulations, and 3) explore the putative proliferative, chemotactic and angiogenic potential of exercise-derived MVs upon human vascular endothelial cells in vitro. Chapter 5 of this thesis describes the time-course of MV appearance in response to prolonged cycling, and demonstrates that intravascular [PMV] increases during and after exercise performed in the heavy intensity domain, whereas [EMV] remains unaltered. Moreover, [PMV] during exercise was related to estimates of vascular shear stress and plasma noradrenaline levels. Results from chapter 6 revealed that PMVs increased in the arterial circulation during passive heat stress, and in the arterial as well as venous circulation during short duration very heavy exercise engaging either a large or small muscle mass. The increases in [PMV] were not directly linked to local changes in vascular shear stress through heat stress and exercise, indicating a systemic PMV response. Finally, chapter 7 revealed that exercise-derived MVs supported endothelial proliferation and migration, while displaying pro-angiogenic potential in vitro. In conclusion, results of this thesis provide original information about MV dynamics, by demonstrating that PMV increase systemically in the circulation not only after but during exercise involving a small and large muscle mass. This MV response seems to be modulated by exercise intensity, and is only partially linked to levels of vascular shear stress. Moreover, circulating MVs produced during exercise present stimulatory angiogenic and mitogenic effects upon endothelial cells in vitro, suggesting a novel potential link between vascular adaptation and exercise training.

Heart failure is the main cause of death for both men and women in the United States. The only proven treatment for patients with heart failure is heart transplantation. The goal of this research is to create patches of tissue that could mimic the function of the native heart to repair the damaged portions of the heart. In this study, whole porcine hearts were decellularized to create a 3D construct that was recellularized with cardiomyocytes (CM) differentiated from human induced pluripotent stem (IPS) cells. At day 4 of differentiation, IPS-derived CMs were implanted onto cardiac extracellular matrix (cECM) and ten days after recellularization, the cells started to beat spontaneously. After implantation, the progenitor CMs continued to proliferate and populate the cECM. A live/dead assay showed the potential of the cECM as a scaffold suitable for recellularization. Confocal microscopy images were taken to evaluate the organization of the cells within the matrix and the impact of the cECM on the growth and maturation of the CMs. Representative cardiac Troponin T (cTNT) and vimentin immunostaining images of CMs derived from iPSCs, on cECM and on standard cell culture plates showed that the cECM allowed the cells to organize and form fibrils with the fibroblasts, compared with CMs cultured in regular culture plates. The timeline of implantation of the cells was a key factor for the development of the heart tissue constructs. Progenitor CMs seeded onto cECM showed better organization and the ability to penetrate 96 µm deep within the collagen fibers and align to them. However, mature CMs seeded onto the matrix showed a disorganized network with very reduced interaction of CMs with fibroblasts, forming two different layers of cells; CMs on top of fibroblasts. In addition, the depth of penetration of the mature CMs within the matrix was only 20 µm. To evaluate the impact of the addition of support cells to the CM monolayer cultures, CMs were co-cultured with human umbilical vein endothelial cells (HUVEC) and it was demonstrated that at ratios of 2:1 HUVEC:CM the beating rate of the CMs was improved from 20 to 112 bpm, additionally, the CM monolayer cultures showed a more synchronized beating pace after the addition of HUVECs. Pharmacological stimulation was performed on CM monolayer cultures using norepinephrine as a stimulator and the results showed that the beating pace of the CMs was improved to 116 bpm after 5 minutes of drug exposure. For future studies, inosculation of the tissue constructs could be performed with the incorporation of membrane proteins to understand the mechanotransduction of the cells. As a preliminary study, the action of dual claudins was evaluated with HUVEC cultures and the results showed the potential of these membrane proteins in the healing of the damaged cell membrane.

Yes / Alterations of Endothelial cells (ECs) play a critical role in different pathogenesis of many serious human diseases, and dysfunction of the vascular endothelium is an indicator for human disorders. Endothelial dysfunction is considered to be an early indicator for atherosclerosis, which is characterised by overexpression of adhesion molecules, including intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1). Hydrogen peroxide (H2O2) released via neutrophils is an important mediator of endothelial cell function. Ambient production of superoxide anion (O2−) and subsequently H2O2 at low levels is critical for regulating endothelial cell functions and proliferation. In this study, we investigated the effects of H2O2 on the expression of adhesion molecules VCAM-1 and ICAM-1 in cultured human umbilical vein endothelial cells (HUVECs). Intracellular superoxide anion production was detected by using p-Nitro Blue Tetrazolium (NBT) assay. Our results showed that administration of 100μM of H2O2 on HUVECs for 2, 6, 12 and 24 h induced a time-dependent increase in ICAM-1 and VCAM-1 mRNA and protein expression levels with a significant increase observed from 6 h. HUVECs exposed to H2O2 exhibit increased O2−, suggesting that H2O2 induced oxidative stress may be a reasonable for atherosclerosis. This increase can be reduced by the flavonoid, N-acetyl cysteine (NAC). The modulation of endothelial cell function through this mechanism may underlie the contribution of H2O2 to the development of vascular disease.

In vitro, la formation de structures de type tubulaire avec des cellules endothéliales de veine ombilicale humaine (HUVEC) a été étudiée en combinant la fonctionnalisation de la chimie des matériaux et le développement de la géométrie tridimensionnelle. Le polycarbonate (PC) a été utilisé comme modèle pour le développement de l'échafaud. Le film de polysaccharide naturel, basé sur un dépôt alternatif couche par couche (LbL) d’acide hyaluronique (HA) et de chitosane (CHI), a d’abord été appliqué sur une surface PC et caractérisé en termes de croissance d’épaisseur microscopie à balayage lascar (CLSM). Cette première fonctionnalisation se traduit par un revêtement complet de la couche PC. Une biofonctionnalisation supplémentaire avec un peptide adhésif (RGD) et deux peptides angiogénétiques (SVV et QK) a été étudiée, immobilisant ces peptides sur le groupe carboxylique de HA précédemment déposé, en utilisant la chimie bien connue du carbodiimide. La version marquée de chaque peptide a été utilisée pour caractériser l’immobilisation et la pénétration des peptides dans les couches de polyélectrolytes, aboutissant à une greffe réussie avec une pénétration complète dans toute l’épaisseur du LbL. Des tests in vitro ont été effectués à l'aide de cellules HUVEC pour évaluer leur efficacité d'adhésion et leur activité métabolique sur la LbL avec et sans immobilisation de peptides, ce qui a permis d'améliorer l'activité préliminaire lorsque des combinaisons de peptides sont utilisées. Enfin, les micro-canaux PC (μCh) ont été développés et caractérisés pour la première fois, et les autres expériences ont été réalisées sur un micromètre de 25 μm de largeur, fonctionnalisé avec une architecture (HA / CHI) 12,5 (PC-LbL) avec des peptides RGD et QK -RGD + QK) ou avec des peptides RGD et SVV (PC-RGD + SVV). Notre première expérience de tubulogénèse a montré de manière surprenante la formation de structures de type tubulaire déjà après 2h d'incubation en utilisant la combinaison double-peptides, mais uniquement avec PC-RGD + QK. Les tubes étaient également présents après 3 et 4 heures de culture. L'expérience de co-culture avec des péricytes humains dérivés du placenta (hPC-PL) montre comment la stabilisation des tubes a été améliorée après 3 et 4 heures également pour l'échantillon de PC-RGD + SVV. Globalement, notre matériel bio-fonctionnel avec les peptides PC-RGD + QK et PC-RGD + SVV permet la formation d'une structure de type tubulaire à la fois dans une expérience de monoculture et de co-culture. / In vitro, tubular-like structures formation with human umbilical vein endothelial cells (HUVECs) was investigated by combining material chemistry functionalization and three-dimensional geometry development. Polycarbonate (PC) was used as a template for the development of the scaffold. Natural polysaccharide’s film based on alternate layer-by-layer (LbL) deposition of hyaluronic acid (HA) and chitosan (CHI), was first applied to PC surface and characterized in terms of thickness growth both, in dry conditions using ellipsometry, and confocal lascar scanning microscopy (CLSM). This first functionalization results in a complete coating of the PC layer. Further biofunctionalization with one adhesive peptide (RGD) and two angiogenetic peptides (SVV and QK) was investigated, immobilizing those peptides on the carboxylic group of HA previously deposited, using the well-known carbodiimide chemistry. The labeled version of each peptide was used to characterize the peptides’ immobilization and penetration into the polyelectrolytes layers, resulting in a successful grafting with complete penetration through the entire thickness of the LbL. In vitro tests were performed using HUVECs to assess their adhesion efficiency and their metabolic activity on the LbL with and without peptide immobilization, resulting in a preliminary improved activity when peptide-combinations is used. Finally, PC micro-channels (μCh) were first developed and characterized, and the rest of the experiments were performed on μCh of 25μm width, functionalized with (HA/CHI)12.5 architecture (PC-LbL) with RGD and QK peptides (PC-RGD+QK) or with RGD and SVV peptides (PC-RGD+SVV). Our first tubulogenesis experiment surprisingly showed the formation of tubular-like structures already after 2h of incubation using the double-peptides combination but only using PC-RGD+QK the tubes were present also after 3 and 4 hours of culture. The co-culture experiment with human pericytes derived from placenta (hPC-PL) demonstrates how the stabilization of the tubes was improved after 3 and 4 hours also for the PC-RGD+SVV sample. Globally our bio-functional material with PC-RGD+QK and PC-RGD+SVV peptides allow the formation of tubular-like structure in both mono and co-culture experiment.

No / Gestational diabetes mellitus (GDM) is known to be associated with fetal endothelial dysfunction, however, the mechanisms are not fully understood. This study examines the effect of maternal diabetes on fetal endothelial function and gene expression under physiological glucose conditions (5 mM). Human umbilical vein endothelial cell (HUVEC) isolated from diabetic mothers (d.HUVEC) grew more slowly than HUVEC isolated from healthy mothers (c.HUVEC) and had delayed doubling time despite increased levels of total vascular endothelial growth factor (VEGF) expression and protein production as determined by real-time PCR and ELISA respectively. Using western blot, the levels of antiproliferative VEGF165b isoform were increased in d.HUVEC relative to c.HUVEC. Successful VEGF165b knockdown by small interfering RNA (siRNA) resulted in increased proliferation of d.HUVEC measured by MTT, compared with negative siRNA control, to similar levels measured in c.HUVEC. In addition, d.HUVEC generated excess levels of ROS as revealed by 2',7' Dichlorodihydrofluorescein Diacetate (DCFH-DA) and Nitrotetrazolium blue (NBT). Using microarray, 102 genes were differentially overexpressed between d.HUVEC versus c.HUVEC (>1.5-fold change; P < 0.05). Functional clustering analysis of these differentially expressed genes revealed participation in inflammatory responses (including adhesion) which may be related to pathological outcomes. Of these genes, ICAM-1 was validated as upregulated, confirming microarray results. Additional confirmatory immunofluorescence staining revealed increased protein expression of ICAM-1 compared with c.HUVEC which was reduced by vitamin C treatment (100 muM). Thus, maternal diabetes induces persistent alterations in fetal endothelial function and gene expression following glucose normalization and antioxidant treatment could help reverse endothelium dysfunction.

Annually, about 80,000 people die in the United Kingdom due to myocardial infarction, congestive heart failure, stroke, or from other diseases related to blood vessels. The current gold standard treatment for replacing the damaged blood vessel is by autograft procedure, during which the internal mammary artery (IMA) graft or saphenous vein graft (SVG) are usually employed. However, some limitations are associated with this type of treatment, such as lack of donor site and post-surgery problems that could negatively affect the patient’s health. Therefore, this present work aims to fabricate a synthetic blood vessel that mimics the natural arteries and to be used as an alternative method for blood vessel replacement. Polymeric materials intended to be used for this purpose must possess several characteristics including: (1) Polymers must be biocompatible; (2) Biodegradable with adequate degradation rate; (3) Must maintain its structural integrity throughout intended use; (4) Must have ideal mechanical properties; and (5) Must encourage and enhance the proliferation of the cells. The feasibility of using synthetic biodegradable polymers such as poly (ε- caprolactone) (PCL) and poly (lactide-co-glycolic acid) (PLGA) for fabricating tubular vascular grafts was extensively investigated in this work. Many fundamental experiments were performed to develop porous tissue- engineered polymeric membranes for vascular graft purposes through electrospinning technique to achieve the main aim. Electrospinning was selected since the scaffolds produced by this method usually resemble structural morphology similar to the extracellular matrix (ECM). Hence, four 6mm in diameter tubular shape vascular grafts PCL only, PLGA only, coaxial (core-PCL and shell-PLGA), and bilayer (inner layer-PCL and outer layer-PLGA) was designed and fabricated successfully. The structure and properties of each scaffold membrane were observed by scanning electron microscopy (SEM), and these scaffolds were fully characterized by Fourier-transform infrared spectroscopy (FT-IR), X-ray diffraction (XRD), thermogravimetric analysis (TGA), water contact angle measurements, mechanical tensile test, as well as cell culture studies were carried out by seeding human umbilical vein cells (HUVEC) and human vascular Fibroblast cells (HVF). Moreover, all polymeric grafts underwent degradation process, and the change in their morphological structure properties was studied over 12 weeks at room temperature. All scaffolds were also exposed to a controlled temperature of 37°C for four weeks, in phosphate-buffered saline solution (pH, 7.3). It was found that all scaffolds displayed exceptional fibre structure and excellent degradability with adequate steady weight-loss confirming the suitability of the fabricated scaffolds for tissue engineering applications. The coaxial and bilayer scaffolds degraded at a much slower (and steadier) rate than the singular PCL and PLGA tubular scaffolds. Coaxial grafts fabricated via coaxial needle showed an increase in their fibre diameter and pore size volume than other membranes, but also showed to have significant tensile strength, elongation at fracture, and Young’s modulus. To conclude, all scaffolds have demonstrated to be reliable to adhere and proliferate HUVEC, and HVF cells, but these cells were attracted to the PLGA membrane more than other fabricated membranes.

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