Human Umbilical Artery Research Articles

Disrupting effects of the emerging contaminant octylmethoxycinnamate (OMC) on human umbilical artery relaxation

Cardiovascular diseases (CVD) represent the number one cause of death worldwide. The vascular endothelium may play a role in the pathophysiology of CVD diseases. Octylmethoxycinnamate (OMC) is a UV-B filter (CAS number: 5466-77-3) widely used worldwide in numerous personal care products, including sunscreens, daily creams, and makeup. This UV-B filter is considered an endocrine disruptor. Therefore, this investigation aimed to evaluate the direct effects of OMC in human umbilical arteries (HUAs) with endothelium and the possible mechanisms involved in the response. The results demonstrated that OMC exerts a rapid (non-genomic) and endothelium-dependent arterial relaxant effect on HUAs previously contracted with serotonin (5-HT) and Histamine (His). On the other hand, when HUAs were contracted with potassium chloride (KCl), the relaxing effect was only observed in HUAs without endothelium, and it appeared to be inhibited in HUAs with endothelium. Thus, the vasorelaxant effect of OMC depends on the endothelium and depends on the contractile agent used, suggesting that OMC may act through different signaling pathways. Furthermore, computational modulation studies, corroborated the binding of OMC to all the proteins under investigation (eNOS, COX-2, ET-1, and TxA2), with higher affinity for COX-2. In summary, the vascular effect of OMC may involve activating different pathways, i.e., acting through the NO pathway, COX pathway, or activating the endothelin-1 pathway.

Vasodilation promoted by (E,E)-farnesol involving ion channels in human umbilical arteries

Background(E,E)-farnesol is a sesquiterpene alcohol derived from plants and animals that exhibits pharmacological properties in the cardiovascular system. However, its effects on human umbilical vessels remain unknown. PurposeThus, this study aims to characterize the vasodilatory effect of (E,E)-farnesol in human umbilical arteries (HUA). Study designThe tissue is obtained from pregnant women over 18 years of age, normotensive, and without prepartum complications. After collected, the tissue was segmented and dissected to remove Wharton's jelly and obtain the umbilical arteries segments. MethodsHUA segments were isolated and sectioned into rings that were subjected to isometric tension recordings in an organ bath. Results(E,E)-farnesol (1 μmol/L to 1 mmol/L) promoted vasodilatory effect in HUA preparations, affecting basal tone, and inhibiting the electromechanical coupling induced by KCl 60 mmol/L with greater potency (EC50 225.3 μmol/L) than the pharmacomechanical coupling induced by 5-HT 10 μmol/L (EC50 363.5 μmol/L). In the absence of extracellular calcium, pharmacomechanical coupling was also abolished, and contractions induced by CaCl2 or BaCl2 were attenuated by (E,E)-farnesol indicating a possible direct inhibition of L-type VOCC as a mechanism of the vasodilatory effect. The vasodilator efficacy of (E,E)-farnesol on reduction of vasocontraction induced by the presence of tetraethylammonium (1 or 10 mmol/L), 4-aminopyridine (1 mmol/L) and glibenclamide (10 μmol/L) suggesting a possible influence of different potassium channels (BKCa, KV and KATP). ConclusionThese results suggest that (E,E)-farnesol may be a promising pharmacological candidate for obstetric hypertensive disorders.

Bioinspired PLCL/Elastin Nanofibrous Vascular Tissue Engineering Scaffold Enhances Endothelial Cells and Inhibits Smooth Muscle Cells.

In vascular tissue engineering, a scaffold that can enhance the proliferation of endothelial cells (ECs) while inhibiting the synthetic differentiation of smooth muscle cells (SMCs) is crucial to prevent thrombus and restenosis after graft implantation. However, it is always challenging to incorporate both properties simultaneously in a vascular tissue engineering scaffold. In this study, a novel composite material was developed by combining a synthetic biopolymer of poly(l-lactide-co-caprolactone) (PLCL) and a natural biopolymer of elastin through electrospinning. Cross-linking of the PLCL/elastin composite fibers using EDC/NHS was performed to stabilize the elastin component. The incorporation of elastin into PLCL was found to enhance the hydrophilicity and biocompatibility of the resulting PLCL/elastin composite fibers, as well as the mechanical properties. Additionally, as a natural component of the extracellular matrix, elastin displayed antithrombotic properties reducing platelet adhesion and improving blood compatibility. Results of cell culture experiments with human umbilical vein ECs (HUVECs) and human umbilical artery SMCs (HUASMCs) showed that the composite fiber membrane had high cell viability, promoting the proliferation and adhesion of HUVECs and inducing a contractile phenotype in HUASMCs. These results indicate that the PLCL/elastin composite material has great potential for use in vascular graft applications due to its favorable properties and rapid endothelialization and contractile phenotypes of cells.

Preparation of biomolecules-mimic poly (ether sulfone)-based hydrogels and their anti-thrombotic, endothelialization and anti-microbial properties

The development of novel hydrogels with high biocompatibility is a vital and active topic of study. Here, we used a facile polycondensation approach to develop multifunctional poly (ether sulfone) (PES)-based hydrogels with diverse functional groups (−SO3Na, −COONa, −OH, and − NH2), as indicated by flourier transformed infrared spectroscopy (FT-IR) and X-ray photoelectron spectroscopy (XPS). The chemical structure of the PES-based hydrogels could be modulated by monomer selection and post-treatment, resulting in a wide range of biomolecule-mimic structures and matching biological functions. In the current investigation, heparin-mimic PES-based hydrogels demonstrated good hemocompatibility, as evident by reduced platelet adhesion and activation, delayed clotting times, suppressed blood-related complement activations, and CD11b expression on leukocytes in the blood. Moreover, hyper-sulfonated PES-based hydrogels showed exceptional cytocompatibility with human umbilical vein endothelial cells (HUVECs) and considerable antiproliferative activity against human umbilical artery smooth muscle cells (HUASMCs), as revealed by cell proliferation and cytotoxicity assay findings. Finally, chitosan-mimic PES-based hydrogels had marked anti-microbial properties against Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus), as shown by the number of colonies. The aforementioned bio-functions were attributed to the formation of various chemical structures on the surfaces of PES-based hydrogels. The distinctive PES-based hydrogels synthesized in this work would be of high utility not only in the synthesis of biomedical hydrogels but also in the rational design of various types of biocompatible materials.

Decellularized Human Umbilical Artery Exhibits Adequate Endothelialization in Xenogenic Transplantation

Decellularized Human Umbilical Artery Exhibits Adequate Endothelialization in Xenogenic Transplantation

The preeclampsia condition alters external potassium-evoked contraction of human umbilical vessels

IntroductionPre-eclampsia (PE) is a disease of high incidence in parturients, that adversely affects both mother and fetus. Although PE prevalence is high, there are few studies on literature describing its etiology or its mechanism of action. Thus, the aim of this study was to elucidate PE-induced alterations of contractile reactivity in umbilical vessels. MethodSegments of human umbilical artery (HUA) and human umbilical vein (HUV) from neonates of normotensive or PE parturients were obtained and contractile responses measured with a myograph. The segments were allowed to stabilize (2 h) under 1.0, 2.0 and 3.0 g force (gf) at pre-stimulation and, then, were stimulated with high isotonic K+ concentrations ([K+]o; 10–120 mM). ResultsAll preparations responded to increases in isotonic K+ concentrations. In HUA and HUV of neonates of normotensive parturients, and in HUV of neonates of PE parturients, the contraction saturated at nearly 50 mM [K+]o, while in HUA of neonates of PE parturients, saturation occurred at 30 mM [K+]o. Additionally, several differences between contractile responses of HUA and HUV from neonates of normotensive parturients and those from neonates of parturients with PE were observed. PE alters the contractile response of the HUA and HUV to increased [K+]o, and its contractile modulation by the pre-stimulus basal tension. Moreover, in HUA of PE, reactivity is decreased for 2.0 and 3.0 gf basal tensions and increased for 1.0 gf; in the HUV of PE condition, it is increased for all basal tensions. DiscussionIn conclusion, PE promotes several alterations in HUA and HUV contractile reactivity, vessels in which important circulatory alterations are known to occur.

Contractile effect of trimethylamine and trimethylamine-n-oxide on isolated human umbilical arteries.

The aim of this study is to investigate the effect of trimethylamine (TMA) and trimethylamine-n-oxide (TMAO) on the contractility of human umbilical artery and the possible mechanisms involved. Vasoactive responses to TMA and TMAO on human umbilical artery rings were measured in isolated organ baths. Cumulative dose-response curves for TMA and TMAO were obtained before and after incubation with atropine, yohimbine, prazosin, indomethacin, verapamil, and Ca+2 -free Krebs-Henselite solution. Administration of cumulative TMA and TMAO resulted in dose-dependent contraction at concentrations ranging from 10 to 100 mM on human umbilical artery rings. TMA-induced contractions were more potent than TMAO-induced contractions (TMA: -logEC50 = 1.00 ± 0.02, TMAO: -logEC50 = 0.57 ± 0.02). Contraction responses to TMA were significantly lower in the presence of verapamil and in the absence of external Ca+2 (p < 0.001, p < 0.05, respectively). Our results showed that TMA and TMAO caused vasoconstriction in isolated human umbilical artery rings. Our findings also indicated that TMA but not TMAO-induced vasoconstriction was partially dependent on extracellular Ca2+ and calcium influx through L-type Ca2+ channels. Our results suggest that TMA and TMAO may have the potential to contribute to cardiovascular diseases through their direct effect on vascular contractility in human arteries.

Decellularized and Genipin Crosslinked Human Umbilical Cord Artery and Vein for Potential Use as Peripheral Nerve Conduit

Critical gap peripheral nerve injury, commonly caused by motor vehicle accidents, results in dysfunctional nerve and impaired body function. Our study aims to develop a conduit from decellularized and genipin crosslinked human umbilical cord artery and vein for future use in critical nerve gap injury treatments. Human umbilical cord arteries (HUCA) and veins (HUCV) were divided into native (nHUCA and nHUCV), decellularized (dHUCA and dHUCV) and genipin-crosslinked (clHUCA and clHUCV) groups. Both the decellularized and crosslinked groups were decellularized, and subsequently, the clHUCA and clHUCV groups were crosslinked with 0.1%, 0.4% and 0.7% (w/v) genipin. The HUCA and HUCV were then studied for decellularization efficiency, crosslinking index, biodegradation, swelling ratio, ultrastructure analysis, flexibility and mechanical strength. In addition, mesenchymal stem cells isolated from Wharton’s jelly were seeded into HUCA and HUCV for biocompatibility studies. The degradation test showed that nHUCV and dHUCV degraded at day 7 compared to other groups that did not show any degradation even after 21 days. Biocompatibility studies showed that the conduits crosslinked with 0.4% (w/v) genipin were successfully seeded and was having the most amount of seeded cells. In conclusion, the decellularization and genipin crosslinking of human umbilical cord artery and vein enabled successful in fabrication of conduit with suitable properties such as reduced swelling, flexibility, porosity and mechanical strength, with potential in tissue engineering applications.

Mechanical characterization of human umbilical arteries by thick-walled models: Enhanced vascular compliance by removing an abluminal lining

The decellularized human umbilical artery (HUA) is considered as a promising option for small-diameter, tissue-engineered vascular grafts (TEVGs). Our previous study showed that the HUA bears a thin, watertight lining on its outermost abluminal surface. Removal of this abluminal lining layer improves efficacy of the perfusion-assisted decellularization of the HUA and increases its compliance. As stress across the wall is believed to affect growth and remodeling of the TEVG, it is imperative to mechanically characterize the HUA using thick-walled models. Combining inflation experiments and computational methods, we investigate the mechanical properties of the HUA before and after the abluminal lining removal to examine the HUA’s wall mechanics. The inflation tests of five HUAs were performed to obtain the mechanical and geometrical response of the vessel wall before and after the lining layer removal. Using nonlinear hyperelastic models, the same responses are obtained computationally using the thick-walled models. The experimental data are incorporated into the computational models to estimate the mechanical and orientation parameters of the fibers and isotropic matrix of different layers in the HUAs. The parameter fitting of both thick-walled models (before and after the abluminal lining removal) results in most of the R-squared values for measuring the goodness of fitting to be over 0.90 for all samples. The compliance of the HUA increases from a mean value of 2.60% per 100 mmHg before the removal of the lining to a mean value of 4.21% per 100 mmHg after the removal. The results reveal that, although the abluminal lining is thin, it is stiff and capable of supporting majority of the high luminal pressure, and that the inner layer is far less stressed than the abluminal lining. Computational simulations also show that removal of the abluminal lining increases the circumferential wall stress by up to 280 kPa under the in vivo luminal pressure. The integrated computational and experimental approaches provide more accurate estimates of the material behaviors of HUAs employed in grafts and, in turn, the study enhances our understanding of interactions between the graft and the native vessel on vascular growth and remodeling.

Evaluation of the bisphenol A-induced vascular toxicity on human umbilical artery

Bisphenol A (BPA) is one of the most widely used synthetic compound in the manufacture of polycarbonate plastics and epoxy resins. Worryingly, BPA is an endocrine disrupting chemical (EDC) with an estrogenic, androgenic, or anti-androgenic activities. However, the vascular implications of BPA exposome in pregnancy is unclear. In this sense, the present work proposed to understand how BPA exposure impair the vasculature of the pregnant women. To elucidate this, ex vivo studies were performed using human umbilical arteries to explore the acute and chronic effects of BPA. The mode of action of BPA was also explored by analysing the activity (by ex vivo studies) and expression (in vitro studies) analysis of Ca2+ and K+-channels and soluble guanyl cyclase. Moreover, in silico docking simulations were performed to unveil the modes of interactions of BPA with the proteins involved in these signalling pathways. Our study showed that the exposure to BPA may modify the vasorelaxant response of HUA, interfering with NO/sGC/cGMP/PKG pathway by modulation of sGC and activation of BKCa channels. Moreover, our findings suggest that BPA can modulate the HUA reactivity, increasing the L-type Ca2+ Channels (LTCC) activity, a common vascular response observed in hypertensive disorders of pregnancy.

Alpha-2-macroglobulin is involved in the occurrence of early-onset pre-eclampsia via its negative impact on uterine spiral artery remodeling and placental angiogenesis

BackgroundPre-eclampsia (PE) is one of the leading causes of maternal and fetal morbidity/mortality during pregnancy, and alpha-2-macroglobulin (A2M) is associated with inflammatory signaling; however, the pathophysiological mechanism by which A2M is involved in PE development is not yet understood.MethodsHuman placenta samples, serum, and corresponding clinical data of the participants were collected to study the pathophysiologic mechanism underlying PE. Pregnant Sprague–Dawley rats were intravenously injected with an adenovirus vector carrying A2M via the tail vein on gestational day (GD) 8.5. Human umbilical artery smooth muscle cells (HUASMCs), human umbilical vein endothelial cells (HUVECs), and HTR-8/SVneo cells were transfected with A2M-expressing adenovirus vectors.ResultsIn this study, we demonstrated that A2M levels were significantly increased in PE patient serum, uterine spiral arteries, and feto-placental vasculature. The A2M-overexpression rat model closely mimicked the characteristics of PE (i.e., hypertension in mid-to-late gestation, histological and ultrastructural signs of renal damage, proteinuria, and fetal growth restriction). Compared to the normal group, A2M overexpression significantly enhanced uterine artery vascular resistance and impaired uterine spiral artery remodeling in both pregnant women with early-onset PE and in pregnant rats. We found that A2M overexpression was positively associated with HUASMC proliferation and negatively correlated with cell apoptosis. In addition, the results demonstrated that transforming growth factor beta 1 (TGFβ1) signaling regulated the effects of A2M on vascular muscle cell proliferation described above. Meanwhile, A2M overexpression regressed rat placental vascularization and reduced the expression of angiogenesis-related genes. In addition, A2M overexpression reduced HUVEC migration, filopodia number/length, and tube formation. Furthermore, HIF-1α expression was positively related to A2M, and the secretion of sFLT-1 and PIGF of placental origin was closely related to PE during pregnancy or A2M overexpression in rats.ConclusionsOur data showed that gestational A2M overexpression can be considered a contributing factor leading to PE, causing detective uterine spiral artery remodeling and aberrant placental vascularization.

Pharmacological Screening of Species from the Lippia genus, Content in Terpenes and Phenylpropanoids, and their Vasorelaxing Effects on Human Umbilical Artery

Naturally occurring bioactive compounds have a plethora of biological effects. In this study, we examined a pharmacological screening of natural products on the human umbilical artery (HUA). HUA preparations were used to follow contractions by KCl (60 mM) and tested at different concentrations (1-5000 μg/mL and μM) of the Lippia alba (EOLa) and Lippia origanoides (EOLo) essential oils, terpenes (citral, limonene perilic alcohol) and phenylpropanoids (eugenol, methyl eugenol). Discussion/Results: The reduction corresponded to approximately 100%, except for limonene (80±1.2 %). When evaluating the concentration of the natural product that promotes 50 % relaxation of the HUA contracted by KCL, EC50 values were: 424.3 μg/mL (EOLa); 468.7±6.7 μg/mL (EOLo); 264.2 ± 8.2 μM (citral); 677.8±5.4 μM (limonene); 186.3±6.4 μM (peryl alcohol); 986.4±7.9 μM (eugenol); and 279.1±4.4 μM (methyl-eugenol). Perillyl alcohol had a lower EC50 (consequently it has a higher pharmacological potency). The plant extracts have a promising vasorelaxing effect in HUAs, paving the way for future investigations: as applications in diseases related to these vessels, such as preeclampsia.

Endothelial PTP4A1 mitigates vascular inflammation via USF1/A20 axis-mediated NF-κB inactivation.

The nuclear factor-κB (NF-κB) signalling pathway plays a critical role in the pathogenesis of multiple vascular diseases. However, in endothelial cells (ECs), the molecular mechanisms responsible for the negative regulation of the NF-κB pathway are poorly understood. In this study, we investigated a novel role for protein tyrosine phosphatase type IVA1 (PTP4A1) in NF-κB signalling in ECs. In human tissues, human umbilical artery ECs, and mouse models for loss of function and gain of function of PTP4A1, we conducted histological analysis, immunostaining, laser-captured microdissection assay, lentiviral infection, small interfering RNA transfection, quantitative real-time PCR and reverse transcription-PCR, as well as luciferase reporter gene and chromatin immunoprecipitation assays. Short hairpin RNA-mediated knockdown of PTP4A1 and overexpression of PTP4A1 in ECs indicated that PTP4A1 is critical for inhibiting the expression of cell adhesion molecules (CAMs). PTP4A1 increased the transcriptional activity of upstream stimulatory factor 1 (USF1) by dephosphorylating its S309 residue and subsequently inducing the transcription of tumour necrosis factor-alpha-induced protein 3 (TNFAIP3/A20) and the inhibition of NF-κB activity. Studies on Ptp4a1 knockout or transgenic mice demonstrated that PTP4A1 potently regulates the interleukin 1β-induced expression of CAMs in vivo. In addition, we verified that PTP4A1 deficiency in apolipoprotein E knockout mice exacerbated high-fat high-cholesterol diet-induced atherogenesis with upregulated expression of CAMs. Our data indicate that PTP4A1 is a novel negative regulator of vascular inflammation by inducing USF1/A20 axis-mediated NF-κB inactivation. Therefore, the expression and/or activation of PTP4A1 in ECs might be useful for the treatment of vascular inflammatory diseases.

Comparison of relaxant effects of nifedipine and NS11021 on isolated umbilical arteries of healthy and preeclamptic pregnant women

ObjectivesPotassium (K+) channel openers and calcium (Ca2+) channel blockers are currently used to treat acute severe hypertension in pregnancy. We aimed to investigate the vasorelaxant effect of NS11021, a potent and specific big-conductance Ca2+-activated K+ (BKCa) channel activator, and to compare it with the vasorelaxant effect of nifedipine on human umbilical arteries (HUAs) isolated from healthy and preeclamptic pregnants. Study designA total of 29 HUAs were isolated immediately after delivery from 14 healthy and 15 preeclamptic pregnant with severe features. The concentration-dependent relaxation responses were obtained to nifedipine and NS11021 on HUAs precontracted with endothelin-1 (ET-1) (10-8 M) in an isolated tissue bath. ResultsBoth nifedipine and NS11021 caused concentration-dependent relaxation responses in HUAs from healthy and preeclamptic pregnants. While the maximum responses (Emax) and pD2 values of nifedipine did not change significantly in both groups, the Emax and pD2 values of NS11021 were significantly decreased in the preeclampsia group (Emax ± SEM; %75.57 ± 4.53 and %43.75 ± 14.00 and pD2 ± SEM; 6.92 ± 0.26 and 5.24 ± 0.53 respectively, p < 0.05). In addition, the pD2 value of NS11021 was not significantly different from that of nifedipine in the control group, but decreased significantly in the preeclampsia group (pD2 ± SEM 7.1 ± 0.41 and 5.2 ± 0.53, p < 0.05, respectively). ConclusionsEfficacy and potency of NS11021 decreased in HUAs from preeclamptic pregnants. Also, NS11021 is less potent than nifedipine in the preeclampsia group. BKCa channels may have a role in the pathogenesis of preeclampsia, however, further experimental studies are needed to elucidate that.

Effects of bisphenol A on human umbilical arteries.

Bisphenol A (BPA) is an endocrine-disrupting chemical widely used in the plastics industry, including food container, toys, and medical equipment. We analyzed the effect of BPA in human umbilical artery contractility and expression of some proteins modulating this function, such as ionic channels and proteins involved in the cGMP pathway. Using standard organ bath technique, rings of human umbilical arteries without endothelium were contracted by 5-HT (1μM) and histamine (10μM) and the effect of different concentrations of BPA (1nM-100μM) was analyzed. The results showed that BPA is a vasodilator of these arteries in a concentration-dependent way. Besides, qPCR studies on human umbilical smooth muscle cells (HUSMC) allowed to analyze the effects of BPA on gene expression. Thus, 12-h exposition to BPA induced reduction of expression of L-type calcium channels (LTCC), alpha subunit of BKCa channels, and Kvβ1 and Kvβ3 from Kv channels. BPA also decreased the expression of soluble guanylate cyclase (sGC) and natriuretic peptide receptor type A (NPRA), meanwhile increasing that of PKG, proteins involved in vasodilation of human umbilical arteries (HUA) by cGMP. Further studies will be necessary to increase knowledge about the implications of these changes induced by BPA exposure.

The Development of Small-Caliber Vascular Grafts Using Human Umbilical Artery: An Evaluation of Methods.

Due to the prevalence of cardiovascular disease in the United States, small-caliber vascular grafts for coronary bypass surgery continue to be in high demand. Human umbilical arteries, an underutilized resource, were decellularized using zwitterionic (3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate [CHAPS]) and ionic (sodium dodecyl sulfate [SDS]) detergents and evaluated as potential vascular grafts. Vessels were tested for decellularization efficacy, mechanical integrity, and recellularization potential. Hematoxylin and eosin staining and DNA quantification revealed moderate to successful removal of cells in both conditions. While CHAPS-decellularized vessels displayed collagen structure most similar to intact tissue, both CHAPS- and SDS-decellularized vessels demonstrated burst pressures lower than that of intact tissue. Alcian Blue staining and sulfated glycosaminoglycan (sGAG) quantification indicated the preservation of sGAG content after both decellularization pathways. Both conditions were also capable of recellularization with human umbilical vein endothelial cells, and the use of a basic fibroblast growth factor treatment did not have a significant effect on the density of adhered cells after 5 days. Whole CHAPS-decellularized vessels were successfully recellularized. Additionally, an evaluation of the effects of freeze-thaw cycles was performed. In summary, human umbilical arteries present a promising alternative for small-caliber vascular grafts due to their high availability and ability to be decellularized and recellularized for safe and successful implantation. Impact Statement Coronary heart disease accounts for one of nine deaths in the United States each year. Bypass surgery has been shown to decrease the risk of heart attack; however, many patients do not have a suitable saphenous vein, which is required to redirect blood flow around their blocked arteries. In this study, we evaluate decellularized umbilical artery as a potential small-diameter vascular graft based on its mechanical properties and its recellularization potential.

PDGF-BB signaling via PDGFR-β regulates the maturation of blood vessels generated upon vasculogenic differentiation of dental pulp stem cells.

A functional vascular network requires that blood vessels are invested by mural cells. We have shown that dental pulp stem cells (DPSC) can undergo vasculogenic differentiation, and that the resulting vessels anastomize with the host vasculature and become functional (blood carrying) vessels. However, the mechanisms underlying the maturation of DPSC-derived blood vessels remains unclear. Here, we performed a series of studies to understand the process of mural cell investment of blood vessels generated upon vasculogenic differentiation of dental pulp stem cells. Primary human DPSC were co-cultured with primary human umbilical artery smooth muscle cells (HUASMC) in 3D gels in presence of vasculogenic differentiation medium. We observed DPSC capillary sprout formation and SMC recruitment, alignment and remodeling that resulted in complex vascular networks. While HUASMC enhanced the number of capillary sprouts and stabilized the capillary network when co-cultured with DPSC, HUASMC by themselves were unable to form capillary sprouts. In vivo, GFP transduced human DPSC seeded in biodegradable scaffolds and transplanted into immunodeficient mice generated functional human blood vessels invested with murine smooth muscle actin (SMA)-positive, GFP-negative cells. Inhibition of PDGFR-β signaling prevented the SMC investment of DPSC-derived capillary sprouts in vitro and of DPSC-derived blood vessels in vivo. In contrast, inhibition of Tie-2 signaling did not have a significant effect on the SMC recruitment in DPSC-derived vascular structures. Collectively, these results demonstrate that PDGF-BB signaling via PDGFR-β regulates the process of maturation (mural investment) of blood vessels generated upon vasculogenic differentiation of human dental pulp stem cells.

Transforming growth factor-β-induced secretion of extracellular vesicles from oral cancer cells evokes endothelial barrier instability via endothelial-mesenchymal transition

BackgroundDuring metastasis, cancer cells undergo epithelial-mesenchymal transition (EMT) in response to transforming growth factor-β (TGF-β), which is abundant in the tumor microenvironment, and acquire invasive and metastatic potentials. Metastasis to distant organs requires intravascular invasion and extravasation of cancer cells, which is accompanied by the disruption of the adhesion between vascular endothelial cells. Cancer cell-derived extracellular vesicles (EVs) have been suggested to induce the destabilization of normal blood vessels at the metastatic sites. However, the roles of EVs secreted from cancer cells that have undergone EMT in the destabilization of blood vessels remain to be elucidated. In the present study, we characterized EVs secreted by oral cancer cells undergoing TGF-β-induced EMT and elucidated their effects on the characteristics of vascular endothelial cells.MethodsInduction of EMT by TGF-β in human oral cancer cells was assessed using quantitative RT-PCR (qRT-PCR) and immunocytochemistry. Oral cancer cell-derived EVs were isolated from the conditioned media of oral cancer cells that were treated with or without TGF-β using ultracentrifugation, and characterized using nanoparticle tracking analysis and immunoblotting. The effects of EVs on human umbilical artery endothelial cells were examined by qRT-PCR, cellular staining, and permeability assay. The significant differences between means were determined using a t-test or one-way analysis of variance with Tukey’s multiple comparisons test.ResultsOral cancer cells underwent EMT in response to TGF-β as revealed by changes in the expression of epithelial and mesenchymal cell markers at both the RNA and protein levels. Oral cancer cells treated with TGF-β showed increased EV production and altered EV composition when compared with untreated cells. The EVs that originated from cells that underwent EMT by TGF-β induced endothelial-mesenchymal transition, which was characterized by the decreased and increased expression of endothelial and mesenchymal cell markers, respectively. EVs derived from oral cancer cells also induced intercellular gap formation which led to the loss of endothelial cell barrier stability.ConclusionsEVs released from oral cancer cells that underwent TGF-β-induced EMT target endothelial cells to induce vascular destabilization. Detailed characterization of oral cancer-derived EVs and factors responsible for EV-mediated vascular instability will lead to the development of agents targeting metastasis.

Growth State-Dependent Expression of Arachidonate Lipoxygenases in the Human Endothelial Cell Line EA.hy926.

Endothelial cells regulate vascular homeostasis through the secretion of various paracrine molecules, including bioactive lipids, but little is known regarding the enzymes responsible for generating these lipids under either physiological or pathophysiological conditions. Arachidonate lipoxygenase (ALOX) expression was therefore investigated in confluent and nonconfluent EA.h926 endothelial cells, which represent the normal quiescent and proliferative states, respectively. mRNAs for ALOX15, ALOX15B, and ALOXE3 were detected in EA.hy926 cells, with the highest levels present in confluent cells compared to nonconfluent cells. In contrast, ALOX5, ALOX12, and ALOX12B mRNAs were not detected. At the protein level, only ALOX15B and ALOXE3 were detected but only in confluent cells. ALOXE3 was also observed in confluent human umbilical artery endothelial cells (HUAEC), indicating that its expression, although previously unreported, may be a general feature of endothelial cells. Exposure to laminar flow further increased ALOXE3 levels in EA.hy926 cells and HUAECs. The evidence obtained in this study indicates that proliferative status and shear stress are both important factors that mediate endothelial ALOX gene expression. The presence of ALOX15B and ALOXE3 exclusively in quiescent human endothelial cells suggests their activity likely contributes to the maintenance of a healthy endothelium.

The Protective Effect of Trichosanthes kirilowii Peel Polysaccharide on the Oxidative Damaged HepG2 and HUASMC Cells.

Background Oxidative stress is an important cause of liver disease and atherosclerosis. Natural substances with antioxidant activity are good drugs for treating liver disease and atherosclerosis. Trichosanthes kirilowii Peel Polysaccharide (TKPP) can remove DPPH (2,2-Diphenyl-1-picrylhydrazyl) free radicals and hydroxyl free radicals in vitro, which shows antioxidant activity. Therefore, it is speculated that it can protect human hepatoma cell line (HepG2) and umbilical artery smooth muscle cell (HUASMC) against oxidative damage by hydrogen peroxide (H2O2). Methods Oxidative damage cell models of HepG2 and HUASMC were induced by H2O2. HepG2 and HUASMC were divided into blank group, H2O2 injury group, TKPP treatment group, and glutathione (GSH) positive control group. Cell Counting Kit-8 (CCK-8) was used to detect cell viability. The level of total GSH and the amount of Nitric oxide (NO) secreted by cells were detected by specific kits. The gene and protein expressions of catalase (CAT) and superoxide dismutase (SOD) were detected by fluorescence quantitative PCR and Western Blot. Results In these two kinds of cells, compared with the control group, the survival rate, total GSH level, and NO secretion, CAT and SOD gene and protein expressions were significantly decreased in the H2O2 damaged group. In the TKPP treatment group, the cell survival rate was significantly elevated with the increase of the polysaccharide concentration, and the total GSH level, NO secretion, CAT and SOD gene expression, and protein expression levels were also significantly increased. Conclusion TKPP can improve the activities of HepG2 and HUASMC cells damaged by H2O2 and protect the cellular antioxidant system.