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The close spatial and temporal connection between osteogenesis and angiogenesis around type H vasculature is referred as "osteogenesis-angiogenesis coupling", which is one of the basic mechanisms of osteogenesis. Endothelial cells (ECs), bone marrow mesenchymal stem cells (BMSCs), and their specific lineage constitute important cluster that participate in the regulation of osteogenesis and angiogenesis in bone microenvironment. However, the regulatory mechanism of osteogenesis-angiogenesis coupling under the condition of bone healing has not been unveiled. In this study, we demonstrated that the exosome derived from ECs (EC-exo) is an initiator of type H blood vessels formation, and EC-exo acts as a mediator in orchestrating osteogenesis-angiogenesis coupling by enhancing BMSC osteogenic differentiation and EC angiogenesis both in monolayer and stereoscopic co-culture system of primary human cells. The transcriptome array indicated that zinc finger and BTB domain containing 16 (ZBTB16) is a key gene in EC-exo-mediated osteogenesis, and ZBTB16 is indispensable in EC-exo-initiated osteogenesis-angiogenesis coupling. Mechanistically, EC-exo up-regulated the expression of ZBTB16 in BMSCs, thereby promoting osteoprogenitor phenotype transformation; the osteoprogenitors further promote ECs which constitute type H vessel (H-ECs) generation by activating HIF-1α pathway; and the H-ECs conversely promotes osteogenic differentiation of BMSCs. The crosstalk between BMSCs and ECs triggered by EC-exo constitutes a positive feedback loop that enhances osteogenesis-angiogenesis coupling. This study demonstrates that EC-exo can become an effective therapeutic tool to promote bone regeneration and repair. [ABSTRACT FROM AUTHOR]

Copyright of Chinese Journal of Contemporary Pediatrics is the property of Xiangya Medical Periodical Press and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Abstract The close spatial and temporal connection between osteogenesis and angiogenesis around type H vasculature is referred as “osteogenesis-angiogenesis coupling”, which is one of the basic mechanisms of osteogenesis. Endothelial cells (ECs), bone marrow mesenchymal stem cells (BMSCs), and their specific lineage constitute important cluster that participate in the regulation of osteogenesis and angiogenesis in bone microenvironment. However, the regulatory mechanism of osteogenesis-angiogenesis coupling under the condition of bone healing has not been unveiled. In this study, we demonstrated that the exosome derived from ECs (EC-exo) is an initiator of type H blood vessels formation, and EC-exo acts as a mediator in orchestrating osteogenesis-angiogenesis coupling by enhancing BMSC osteogenic differentiation and EC angiogenesis both in monolayer and stereoscopic co-culture system of primary human cells. The transcriptome array indicated that zinc finger and BTB domain containing 16 (ZBTB16) is a key gene in EC-exo-mediated osteogenesis, and ZBTB16 is indispensable in EC-exo-initiated osteogenesis-angiogenesis coupling. Mechanistically, EC-exo up-regulated the expression of ZBTB16 in BMSCs, thereby promoting osteoprogenitor phenotype transformation; the osteoprogenitors further promote ECs which constitute type H vessel (H-ECs) generation by activating HIF-1α pathway; and the H-ECs conversely promotes osteogenic differentiation of BMSCs. The crosstalk between BMSCs and ECs triggered by EC-exo constitutes a positive feedback loop that enhances osteogenesis-angiogenesis coupling. This study demonstrates that EC-exo can become an effective therapeutic tool to promote bone regeneration and repair. Graphical Abstract

BACKGROUND: Increased homocysteine level induces apoptosis of human umbilical vein endothelial cells, but the mechanism remains unclear. OBJECTIVE: To investigate the role of hsa-circ-0001360 in human umbilical vein endothelial cell apoptosis induced by homocysteine. METHODS: In vitro cultured human umbilical vein endothelial cells were divided into control group, homocysteine group, interference control group, interference control + homocysteine group, hsa-circ-0001360 interference group, hsa-circ-0001360 + homocysteine interference group, overexpression control group, overexpression control + homocysteine group, hsa-circ-0001360 overexpression group and hsa-circ-0001360 + homocysteine overexpression group. All groups were treated with 100 μmol/L homocysteine. After 72 hours of intervention, the expressions of apoptosis-related proteins Bax, Bcl-2, and Caspase-3 were detected by western blot assay. The apoptotic rate was detected by flow cytometry. Quantitative real-time PCR was used to detect the expression of hsacirc-0001360. RESULTS AND CONCLUSION: (1) Compared with the control group, the expression of Caspase-3 and Bax was significantly increased (P < 0.01), and the expression of Bcl-2 was significantly decreased (P < 0.01), and the apoptotic rate was significantly increased (P < 0.01) in the homocysteine group. (2) Compared with control group, the expression of hsa-circ-0001360 was significantly increased in the homocysteine group (P < 0.01). (3) The expression of hsa-circ-0001360 was significantly higher in the cytoplasm than that in the nucleus (P < 0.01). (4) Compared with the interference control C group and interference control + homocysteine group, the expressions of Caspase-3 and Bax were significantly decreased (P < 0.01), while the expression of Bcl-2 was significantly increased (P < 0.01); the apoptotic rate was significantly decreased (P < 0.01) in sh-hsa-circ-0001360 interference group and sh-hsa-circ-0001360 + homocysteine interference group. (5) Compared with overexpression control group and overexpression control + homocysteine group, the expressions of Caspase-3 and Bax were significantly increased (P < 0.01), while the expression of Bcl-2 was significantly decreased (P < 0.01); the apoptotic rate was significantly increased (P < 0.01) in the hsa-circ-0001360 overexpression group and the hsa-circ-0001360 + homocysteine overexpression group. (6) In conclusion, hsa-circ-0001360 can promote the apoptosis of human umbilical vein endothelial cells induced by homocysteine. [ABSTRACT FROM AUTHOR]

BACKGROUND: Combining seed cells with 3D bioprinting technology enables the specific construction of various tissues and organs to meet the demands of tissue repair. However, further research is needed on the promotion of angiogenesis in damaged tissues. OBJECTIVE: By cultivating a 3D scaffold structure of methacrylated gelatin loaded with fibroblasts, obtaining the supernatant, and mixing it in different proportions with a complete culture medium to simulate the cellular microenvironment during tissue repair, this study aimed to explore the role of various cellular microenvironments in promoting angiogenesis in endothelial cells. METHODS: A methacrylated gelatin scaffold structure loaded with fibroblasts was prepared using an extrusion-based 3D bioprinting process. Hydrogel scaffold extract was prepared and mixed with a complete culture medium in ratios of 1:1, 1:2, and 1:4 to obtain conditioned medium. Mouse embryonic fibroblasts BALB3T3 and human umbilical vein endothelial cells were co-cultured with complete medium (control group) and hydrogel scaffold extract, respectively. Cell proliferation was assessed using the CCK-8 assay and cell viability was analyzed using live/dead staining. Three kinds of conditioned medium and complete medium (control group) were used to co-culture with human umbilical vein endothelial cells for tube formulation assay, vascular genetic testing, and immunofluorescence staining of CD31. RESULTS AND CONCLUSION: (1) Scanning electron microscopy revealed that the methacrylated gelatin scaffold exhibited a porous structure, and rheological results demonstrated excellent mechanical properties of the hydrogel. CCK-8 assay and live/dead cell staining showed that the hydrogel scaffold extract had no obvious cytotoxicity. (2) Tube formulation assay indicated that the hydrogel showed the total length of cell tubules in 1:1 conditioned medium group was smaller than that in the control group (P < 0.05). There were no statistical differences among the four groups in the number of vascular branches formed by endothelial cells (P > 0.05). (3) qRT-PCR results showed that for vascular endothelial growth factor mRNA expression, the 1:2 conditioned medium group was lower than the 1:1 conditioned medium group on day 1 (P < 0.01). On day 3, the expression level of vascular endothelial growth factor in the 1:2 conditioned medium group was higher than that in the control group (P < 0.01). On day 5, the cytokine expression level in the 1:2 conditioned medium group was significantly higher than that in the other three groups (P < 0.01 or P < 0.000 1). The expression in the 1:1 conditioned medium group was significantly lower than that in the other three groups (P < 0.05 or P < 0.01). On day 1, the expression level of basic fibroblast growth factor in the 1:1 conditioned medium group was significantly higher than that in the control group and 1:4 conditioned medium group (P < 0.01, P < 0.05). The expression was higher in the 1:2 conditioned medium group than that in the control group (P < 0.05). On day 3, the expression levels of cytokines in the 1:4 conditioned medium group was higher than that in the control group (P < 0.05). (4) On day 3, the expression of CD31 in the 1:2 conditioned medium group was higher than that in the control group and the 1:4 conditioned medium group (P < 0.05). (5) The results indicate that the resulting conditioned media can simulate the microenvironment of vascular regeneration after tissue damage, promoting the vascularization process of endothelial cells. The best promotion of vascularization in endothelial cells was observed when the ratio of supernatant to complete culture medium was 1:2. [ABSTRACT FROM AUTHOR]

Refractory diabetic wounds are still a clinical challenge that can cause persistent inflammation and delayed healing. Exosomes of adipose stem cells (ADSC-exos) are the potential strategy for wound repair; however, underlying mechanisms remain mysterious. In this study, we isolated ADSC-exos and identified their characterization. High glucose (HG) stimulated human umbilical vein endothelial cells (HUVECs) to establish in vitro model. The biological behaviors were analyzed by Transwell, wound healing, and tube formation assays. The underlying mechanisms were analyzed using quantitative real-time PCR, co-immunoprecipitation (Co-IP), IP, and western blot. The results showed that ADSC-exos promoted HG-inhibited cell migration and angiogenesis. In addition, ADSC-exos increased the levels of TRIM32 in HG-treated HUVECs, which promoted the ubiquitination of STING and downregulated STING protein levels. Rescue experiments affirmed that ADSC-exos promoted migration and angiogenesis of HG-treated HUVECs by regulating the TRIM32/STING axis. In conclusion, ADSC-exos increased the levels of TRIM32, which interacted with STING and promoted its ubiquitination, downregulating STING levels, thus promoting migration and angiogenesis of HG-treated HUVECs. The findings suggested that ADSC-exos could promote diabetic wound healing and demonstrated a new mechanism of ADSC-exos. [ABSTRACT FROM AUTHOR]

Objective: To discuss the effect of sodium tanshinone ⅡA sulfonate (STS) on the function of human umbilical vein endothelial cells (hUVECs) after treated with uremic toxin, and to clarify its mechanism. Methods: The hUVECs were passaged and divided into blank control group, uremic toxinstimulation group, uremic toxin + STS group, and uremic toxin + STS + extracellular signal-regulated kinase (ERK) inhibitor group. The concentration of STS used in the last two groups was 10 mg.L-1. The shear stress stimulation at 12 dyn.cm-2 was applied to the cells in various groups. The proliferation activities of the cells in various groups were detected by CCK-8 assay; the expression levels of ERK, nuclear factor kappa B (NF-κB), and type Ⅰ collagen proteins in the cells in various groups were detected by Western blotting method; the expression levels of ERK, NF- κB, and type Ⅰ collagen mRNA in the cells in various groups were detected by real-time fluorescence quantitative PCR (RT-qPCR) method; the apoptotic rates the cells in various groups were detected by TUNEL method. Results: The CCK-8 assay results showed that after treated with shear stress, the probiferation activitres of the cells in uremic toxin-stimulation group and uremic toxin + STS + ERK inhibitor group were lower than that in uremic toxin + STS group (P<0. 01). The Western blotting results showed that compared with uremic toxin group, the expression levels of ERK, NF-κB, and type Ⅰ collagen proteins in the cells in uremic toxin + STS group were increased (P<0. 01). After inhibiting the ERK pathway, compared with blank control group, uremic toxin group, and uremic toxin + STS group, the expression levels of ERK, NF- κB, and type Ⅰ collagen proteins in the cells in uremic toxin + STS + ERK inhibitor group were significantly decreased (P<0. 01). The RT-qPCR results showed that compared with uremic toxin group, the expression levels of ERK, NF-κB, and type Ⅰ collagen mRNA in the cells in uremic toxin + STS group were increased (P<0. 01). After inhibiting the ERK signaling pathway, compared with blank control group, uremic toxin group, and uremic toxin + STS group, the expression levels of ERK, NF- κB, and type Ⅰ collagen mRNA in the cells in uremic toxin + STS + ERK inhibitor group were significantly decreased (P<0. 01). The TUNEL method detection results showed that the apoptotic rate in the cells in uremic toxin + STS group was lower than those in uremic toxin-stimulation group and uremic toxin + STS + ERK inhibitor group (P<0. 05). Conclusion: A certain concentration of STS can improve the proliferation of the endothelial cells and reduce the apoptosis of the cells after treated with uremic toxins by modulating the expressions of NF- κB and type Ⅰ collagen mRNA and proteins through the ERK signaling pathway. [ABSTRACT FROM AUTHOR]

Inward rectifier K+ (Kir) channel, a major factor determining endothelial membrane potential, regulates Ca2+ influx and vasodilator release, which is impaired in preeclamptic blood vessels. Previously, human umbilical vein endothelial cell (HUVEC) Kir currents were shown to decrease after incubating in preeclamptic plasma. We aimed to demonstrate whether sFlt-1, which is high in preeclamptic blood, could inhibit Kir channel function and expression. HUVECs were cultured in regular medium, regular medium with added sFlt-1, or serum from preeclampsia patients or normal pregnant women (Control, sFlt-1, PE, or NP, respectively). Using whole-cell patch clamp technique, we identified Kir currents with the Kir blocker 2 mM BaCl 2 and compared the currents among groups. The expression of Kir 2.1 and 2.2 channels were determined using immunofluorescent staining. sFlt-1 and PE groups exhibited similar Kir currents, while NP group possessed significantly larger currents, similar to Control group currents. Moreover, sFlt-1 and sFlt-1/PlGF ratio showed strong negative correlation with Kir currents (r = −0.71 and −0.70, respectively; P < 0.05). There were no significant differences in mean fluorescence intensity representing Kir 2.1 and 2.2 channels expression in all four groups. This is the first report to demonstrate sFlt-1 inhibition against Kir currents, which could lead to maternal endothelial dysfunction and hypertension seen in preeclampsia. However, channel expression was unaffected by sFlt-1 incubation, suggesting dysfunctions of channel or other processes (e.g., membrane translocation). The present data could pave the way for novel therapies targeting sFlt-1 or Kir to alleviate hypertension in preeclampsia. • HUVEC Kir currents were inhibited by sFlt-1 or preeclampsia (PE) serum incubation. • Kir current amplitude and sFlt-1 concentration were negatively correlated. • No change in Kir expression after incubating with sFlt-1 or PE serum. • The link between Kir and sFlt-1 may explain hypertension in PE. [ABSTRACT FROM AUTHOR]

BACKGROUND: Dapagliflozin, an inhibitor of sodium-glucose cotransporter 2, can delay the progression of atherosclerosis by regulating glucose metabolism, inhibiting inflammation and improving endothelial cell function. OBJECTIVE: To study the effect of dapagliflozin on cell pyroptosis and endothelial dysfunction induced by oxidized low-density lipoprotein. METHODS: Human umbilical vein endothelial cells were divided into a control group (no intervention), a model group (treated with oxidized low-density lipoprotein for 24 hours), and a dapagliflozin group (treated with oxidized low-density lipoprotein + dapagliflozin for 24 hours). Endothelial cell proliferation activity was measured by cell counting kit-8 assay. The levels of intercellular adhesion molecule 1, vascular cell adhesion molecule 1, and monocyte chemotactic protein-1 in cell supernatant were detected using ELISA. Nitric oxide level in the cells was detected by nitrate reductase assay. The pyroptosis rate and characteristics of endothelial cells were detected by Hoechst 33342/PI fluorescence co-staining and lactate dehydrogenase release assay. The protein expression levels of NLRP3, caspase-1, GSDMD, interleukin-1β, and interleukin-18 were detected by western blot assay. RESULTS AND CONCLUSION: (1) Oxidized low-density lipoprotein could cause pyroptosis and dysfunction of endothelial cells. (2) Compared with the control group, the level of nitric oxide and cell activity were decreased (P < 0.05), while lactate dehydrogenase, intercellular adhesion molecule 1, vascular cell adhesion molecule 1, and monocyte chemotactic protein-1 levels were significantly increased in the model group (P < 0.05). Compared with the model group, cell activity and nitric oxide levels significantly increased (P < 0.05), but lactate dehydrogenase, intercellular adhesion molecule 1, vascular cell adhesion molecule 1, and monocyte chemotactic protein-1 levels were significantly diminished in the dapagliflozin group (P < 0.05). (3) Compared with the model group, cell pyroptosis rate and the protein expression of pyroptosis factor NLRP3, caspase-1, GSDMD, interleukin-18 and interleukin-1β significantly reduced in the dapagliflozin group (P < 0.05). (4) The results indicate that dapagliflozin inhibits oxidized low-density lipoprotein-induced endothelial pyroptosis and ameliorates endothelial cell dysfunction. [ABSTRACT FROM AUTHOR]

Background G protein-coupled receptor 40 (GPR40) is a key molecule in diabetes and fatty liver, but its role in endothelial dysfunction remains unclear. Our objective in this study was to determine whether GPR40 agonists protect endothelial cells against palmitatemediated oxidative stress. Methods Human umbilical vein endothelial cells (HUVECs) were used to investigate effects of various GPR40 agonists on vascular endothelium. Results In HUVECs, AM1638, a GPR40-full agonist, enhanced nuclear factor erythroid 2–related factor 2 (NRF2) translocation to the nucleus and heme oxygenase-1 (HO-1) expression, which blocked palmitate-induced superoxide production. Those antioxidant effects were not detected after treatment with LY2922470 or TAK875, GPR40-partial agonists, suggesting that GPR40 regulates reactive oxygen species (ROS) removal in a ligand-dependent manner. We also found that palmitate-induced CCAAT/enhancer‐binding protein homologous protein expression; X-box binding protein-1 splicing, nuclear condensation, and fragmentation; and caspase-3 cleavage were all blocked in an NRF2-dependent manner after AM1638 treatment. Both LY2922470 and TAK875 also improved cell viability independent of the NRF2/ROS pathway by reducing palmitate-mediated endoplasmic reticulum stress and nuclear damage. GPR40 agonists thus have beneficial effects against palmitate in HUVECs. In particular, AM1638 reduced palmitate-induced superoxide production and cytotoxicity in an NRF2/HO-1 dependent manner. Conclusion GPR40 could be developed as a good therapeutic target to prevent or treat cardiovascular diseases such as atherosclerosis.

After skin injury, wound repair involves a complex process in which angiogenesis plays a crucial role. Previous research has indicated that fucoidan may aid in wound healing; we therefore hypothesised that fucoidan may speed up the process by promoting angiogenesis. In this study, we investigated the potential molecular mechanism underlying fucoidan's ability to accelerate wound healing by promoting angiogenesis. Using a full‐cut wound model, we observed that fucoidan significantly intensified wound closure and promoted granulation formation and collagen deposition. Immunofluorescence staining revealed that fucoidan also promoted wound angiogenesis, specifically by accelerating the migration of new blood vessels to the middle area of the wound. Furthermore, fucoidan demonstrated the ability to enhance the proliferation of human umbilical vein endothelial cells (HUVECs) damaged by hydrogen peroxide (H2O2) and to improve the formation of endothelial tubes. Mechanistic studies revealed that fucoidan upregulated the protein levels of the AKT/Nrf2/HIF‐1α signalling pathway, which plays a crucial role in angiogenesis. This was further confirmed using the inhibitor LY294002, which reversed the promotion of endothelial tube formation by fucoidan. Overall, our findings suggest that fucoidan can promote angiogenesis via the AKT/Nrf2/HIF‐1α signalling pathway and accelerate wound healing. [ABSTRACT FROM AUTHOR]

BACKGROUND: Vascular endothelial cells are the main target of arsenic toxicity and the molecular mechanism of arsenic-induced endothelial cell injury needsto be further studied. OBJECTIVE: To study the effects of sodium arsenite on human umbilical vein endothelial cell injury and sphingosine kinase 1/sphingosine 1-phosphate signaling axis of human umbilical vein endothelial cells. METHODS: Human umbilical vein endothelial cells were isolated, cultured, and identified. The cells were treated with 0, 5, 10, 15, 20, 25 µmol/L sodium arsenite for 24 hours. Cell viability was detected by cell counting kit-8. Cell morphology was observed by inverted phase contrast microscope. The content of intracellular reactive oxygen species was detected by fluorescent probe DCFH-DA. Annexin V-FITC/PI double labeling combined with flow cytometry and TUNEL were used to detect apoptosis. The content of sphingosine 1-phosphate in cells was detected by ELISA. The mRNA and protein expressions of vascular cell adhesion molecule-1, sphingosine kinase 1, and sphingosine 1-phosphate were detected by real-time fluorescence quantitative PCR and western blot respectively. RESULTS AND CONCLUSION: Compared with the control group (0 µmol/L sodium arsenite), the cell viability decreased gradually along with the increased concentration of sodium arsenite in a dose-dependent manner; the cell fusion rate decreased gradually, the cell gap widened gradually, and the number of exfoliated and floating cells increased gradually; the reactive oxygen species content and apoptosis rate increased gradually; the sphingosine 1-phosphate content in cells increased gradually; the relative mRNA and protein expressions of vascular cell adhesion molecule-1 and sphingosine kinase 1 increased gradually, but the relative mRNA and protein expression of sphingosine kinase 1 decreased gradually. To conclude, sodium arsenite-injured human umbilical vein endothelial cell injury may be related to the activation of sphingosine kinase 1/sphingosine 1-phosphate signaling axis and the downregulation of sphingosine 1-phosphate receptor 1. Sphingosine kinase 1/sphingosine 1-phosphate signaling axis may become a new target of arsenic-induced cardiovascular injury. [ABSTRACT FROM AUTHOR]

Analytical screening and validation systems based on a combination of cell membrane chromatography and two‐dimensional chromatography‐tandem mass spectrometry are incapable of providing prepared samples containing the active ingredients found in traditional Chinese medicine; therefore, these samples cannot be directly used in subsequent studies. In this study, a semi‐preparative cell membrane chromatography column was developed using a hydrogel‐modified carrier and human umbilical vein endothelial cells to optimize prepared conditions, such as hydrogel polymerization, cell fragmentation, and cell membrane volume. This increased the binding ratio of membrane protein and carrier to 15.79 mg/g. The column was systematically evaluated using multitarget tyrosine kinase inhibitors that displayed good specificity and reproducibility. Subsequently, using the column coupled with a semi‐preparative high‐performance liquid chromatography‐offline‐high‐performance liquid chromatography‐mass spectrometry system, 15 active ingredients were screened and purified from Indigo naturalis, and five main components were identified: l‐lysine, oxyresveratrol, tryptanthrin, isorhamnetin, and indirubin. Furthermore, the pharmacological effects of the ingredients were confirmed using cell proliferation and apoptosis assays. Results revealed potent proliferation‐inhibiting and apoptosis‐promoting abilities on human chronic myelogenous leukemic cells and human promyelocytic leukemic cells (p < 0.001). Overall, the system presented screening and purification functions that could be used to prepare I. naturalis samples acting on the epidermal growth factor receptor and vascular endothelial cell growth factor. [ABSTRACT FROM AUTHOR]

BACKGROUND: High density polyethylene has been widely used as a repair material for cranial and maxillofacial bone defects, but its preparation method and surface activity still need further improvement. OBJECTIVE: To optimize the preparation method of high density polyethylene and improve the surface activity of high density polyethylene. METHODS: The high density polyethylene scaffolds were prepared by extrusion 3D printing technology, and the scaffolds were immersed in dopamine solution and simulated body fluid successively to be coated with polydopamine and hydroxyapatite. The microstructure, hydrophilicity and compression modulus of the scaffolds before and after coating were characterized. Mouse embryonic osteogenic precursor cells (MC3T3-E1) and human umbilical vein endothelial cells were inoculated on the surface of the scaffold to evaluate the cytocompatibility and early osteogenic and angiogenic differentiation of the scaffold before and after coating. RESULTS AND CONCLUSION: (1) The 3D printed high density polyethylene scaffold’s fibers were arranged regularly and pores were uniform. Characterization results showed that polydopamine and hydroxyapatite coatings were successful on the surface of the scaffolds. Compared with the unmodified scaffold, the surface water contact angle of the scaffold modified with polydopamine coating and polydopamine + hydroxyapatite coating decreased significantly (P < 0.05), and the compressive modulus did not change significantly. (2) Compared with the unmodified scaffold, the modified scaffold with polydopamine coating and polydopamine + hydroxyapatite coating could promote the adhesion of MC3T3-E1 cells and human umbilical vein endothelial cells (P < 0.05). The cell adhesion promoting effect of double coating modified group was better than that of single coating modified group (P < 0.05). Compared with the unmodified scaffold, the modified scaffold with polydopamine coating and polydopamine + hydroxyapatite coating could promote the proliferation of MC3T3-E1 cells and human umbilical vein endothelial cells (P < 0.05). The proliferation promoting effect of double coating modified group was better than that of single coating modified group (P < 0.05). Compared with unmodified scaffolds, the scaffolds modified with polydopamine coating and polydopamine + hydroxyapatite coating could promote the osteogenic differentiation of MC3T3-E1 cells (P < 0.05), and increase the expression of angiogenic factor CD31 in human umbilical vein endothelial cells. The double-coating modified group was more obvious. (3) The results showed that the high density polyethylene scaffolds coated with polydopamine and hydroxyapatite based on 3D printing technology have good cytocompatibility and early osteogenic and angiogenic differentiation ability. [ABSTRACT FROM AUTHOR]

In order to provide a biomimetic natural extracellular matrix microenvironment with excellent mechanical capacity for tissue regeneration, a novel porous hybrid glycidyl methacrylate-modified silk fibroin/poly(L-lactic acid-ε-caprolactone)-polyethylene glycol diacrylate (SFMA/P(LLA-CL)-PEGDA) hybrid three-dimensional (3D) nanofibrous scaffolds was successfully fabricated through the combination of 3D nanofibrous platforms and divinyl PEGDA based photocrosslinking, and then further improved water resistance by ethanol vapor post-treatment. Scanning electron microscopy and micro-computed tomography results demonstrated significant PEGDA hydrogel-like matrices bonded nanofibers, which formed a 3D structure similar to that of “steel bar (nanofibers)–cement (PEGDA)”, with proper pore size, high porosity, and high pore connectivity density. Meanwhile, the hybrid 3D nanofibrous scaffolds showed outstanding swelling properties as well as improved compressive and tensile properties. Furthermore, these hybrid 3D nanofibrous scaffolds could provide a biocompatible microenvironment, capable of inducing the material–cell hybrid and regulating human umbilical vein endothelial cells proliferation. They thus present significant potential in tissue regeneration. [ABSTRACT FROM AUTHOR]

Copyright of Chinese Journal of Contemporary Pediatrics is the property of Xiangya Medical Periodical Press and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

This study investigates the effects of simvastatin (SMV) on human umbilical vein endothelial cells (HUVECs) inflamed by tumor necrosis factor-alpha (TNF-a). Cells were treated with simvastatin (5 µM, 10 µM, and 50 µM) with/without TNF (20 ng/mL). The effects of simvastatin on the proliferation of cells were examined, and its cytotoxic effect was determined by lactate dehydrogenase (LDH) assays. Metalloproteinase expression (MMP-2 and MMP-9) levels of SMV at different concentrations were determined by qPCR, and protein levels were determined by Western blot. The increasing concentrations of SMV down-regulated the expression levels of MMPs. After stimulation with TNF-a, the effect of SMV at different concentrations on wound healing time in case of wound formation was examined. The results interestingly indicated that SMV delayed the closure time 2.5-fold when it was administered alone to HUVECs but did not change the wound closure time in inflamed conditions. The study suggests that inflammation is a required condition in wound healing, and in the absence of inflammation, SMV treatment delays wound healing in vitro. [ABSTRACT FROM AUTHOR]

ABSTRACT The Treponema pallidum membrane protein Tp47 induces immunocyte adherence to vascular cells and contributes to vascular inflammation. However, it is unclear whether microvesicles are functional inflammatory mediators between vascular cells and immunocytes. Microvesicles that were isolated from Tp47-treated THP-1 cells using differential centrifugation were subjected to adherence assays to determine the adhesion-promoting effect on human umbilical vein endothelial cells (HUVECs). Intercellular adhesion molecule 1 (ICAM-1) and vascular cell adhesion molecule 1 (VCAM-1) levels in Tp47-induced microvesicle (Tp47-microvesicle)-treated HUVECs were measured, and the related intracellular signaling pathways of Tp47-microvesicle-induced monocyte adhesion were investigated. Tp47-microvesicles promoted THP-1 cell adhesion to HUVECs (P

In this study low-input RNA-sequencing was used to annotate the molecular identity of endothelial cells isolated and immunopurified with CD144 microbeads. Using this technique, comparative gene expression profiling from healthy subjects and patients with type 2 diabetes mellitus identified both known and novel pathways linked with EC dysfunction. Modeling of diabetes by treating cultured ECs with high glucose identified shared changes in gene expression in diabetic cells. Overall, the data demonstrate how purified ECs from patients can be used to generate new hypotheses about mechanisms of human vascular disease.

Autophagy has been extensively studied and occurs in many biological settings. However, a question remains as to whether ischemia enhances Beclin-1/LC3-II-dependent macroautophagy in vascular endothelial cells, as has been previously thought. Furthermore, the effect of the level of autophagy on cell or skin flap survival still requires elucidation. We created a lethal ischemia model in human umbilical vascular endothelial cells (HUVECs), performed quantitative proteomics and bioinformatics analyses, and verified the autophagic status and effect both in vitro and in vivo. The significantly upregulated proteins encoded by autophagy-related genes (ATGs) included ATG2A, ATG3, ATG4B, ATG5, ATG7, ATG9A, ATG12, ATG16, and ATG101. The significantly enhanced lysosomal proteins were cathepsin B, cathepsin D, lysosome-associated membrane protein 1 (LAMP1), and LAMP2. However, the differentially expressed proteins excluded Beclin-1, microtubule-associated protein light chain 3 (LC3)-I, and LC3-II. Western blot analyses verified that the protein expression levels of Beclin-1, LC3-I, and LC3-II were neither upregulated nor downregulated in ischemia-challenged HUVECs. The autophagic status was not enhanced by rapamycin in ischemic HUVECs but appeared to be inhibited by chloroquine. Our in vivo study on rats showed that a downregulation in autophagic status jeopardized skin flap survival. In conclusion, Ischemia neither enhanced nor inhibited Beclin-1/LC3-II-dependent canonical macroautophagy both in vitro and in vivo, in contradiction to previous studies. An appropriate autophagic homeostasis can minimize cell or skin flap damage.

Objective：To investigate the effect of lncRNA-MIAT on M2-type polarization of tumorassociated macrophages, and to elucidate its possible mechanism. Methods：The THP-1 cells were cultured in vitro. The lentivirus particles, which over expressed (LV-MIAT) and down regulated expression of lncRNA-MIAT (LV-shMIAT), and empty vector (LV-Vector) were transfected into the THP-1 cells. The THP-1 cells were activated into the macrophages, and the activated macrophages were co-cultured with the osteosarcoma (OS)MG63 cells by using Transwell co-culture system. The above co-culture systems were divided into MG63+LV-Vector group (positive control group), MG63+LVshMIAT group, MG63+LV-MIAT group, and IL-4+LV-Vector group. The expression levels of lncRNA-MIAT in the THP-1 cells in various groups were detected；the percentages of M2-type macrophages in various groups were detected by flow cytometry；the levels of vascular endothelial growth factor (VEGF), interleukin-10 (IL-10), and transforming growth factor-β1 (TGF-β1)in supernatant of the THP-1 cells in various groups were detected by ELISA method；the macrophages in each culture system were co-cultured with the human umbilical vein endothelial cell (HUVEC)；EDU staining was used to detect the proliferation activities of the HUVEC in various groups；the numbers of angiogenesis in the HUVEC in various groups were detected by tube formation assay；the expression levels of Janus kinase 1 (JAK1), signal transducer and activator of transcription 6 (STAT6), and phosphorylated STAT6 proteins in the THP1 cells and the expression levels of vascular endothelial growth factor receptor 2 (VEGFR2), Notch1, and delta like protein 4 (DLL4)in the HUVEC in various groups were detected by Western blotting method；the p-STAT6/STAT6 ratio was calculated. The OS tumor-bearing mouse model was constructed, and 36 nude mice were divided into LV-Vector group, LV-shMIAT group, and LV-MIAT group (n=12). The volumes and weights of the tumor and the positive expression rates of CD163 and CD31 in tumor tissue of the mice in various groups were detected after interfering lncRNA-MIAT expression. Results：The THP-1 cells with stable over-expression or down-regulation of lncRNA-MIAT were successfully established by lentivirus infection. Compared with LV-Vector group, the expression level of lncRNA MIAT in the cells in LV-shMIAT group was significantly decreased (P<0. 05), and the expression level of lncRNA MIAT in the cells in LV-MIAT group was significantly increased (P<0. 05). Compared with MG63+LV-Vector group, the levels of VEGF, IL-10, and TGF- β1 in the THP-1 cells in MG63+LV-shMIAT group were significantly decreased (P<0. 05), and the expression levels of VEGFR2, Notch1, and DLL4 proteins in the HUVEC were significantly decreased (P<0. 05), the levels of VEGF, IL-10, and TNF- β1 in the THP-1 cells in MG63+LV-MIAT and IL-4+LV-Vector groups were increased significantly (P<0. 05 or P<0. 01)；the expression levels of VEGFR2, Notch1, and DLL4 proteins in HUVEC were increased (P<0. 05), the percentage of M2-type macrophages, and p-STAT6/ STAT6 ratio and expression level of JAK1 protein in the THP-1 cells were increased (P<0. 05)；compared with MG63+sh-MIAT group, the VEGF level in the THP-1 cells in MG63+LV MIAT group was decreased significantly (P<0. 05), the levels of IL-10 and TGF-β were decreased significantly (P<0. 05), the expression levels of VEGFR2, Notch1, and DLL4 proteins in the HUVEC were increased significantly (P<0. 05)；compared with MG63+LV-Vector group, the proliferation activity and number of angiogenesis in the HUVEC in MG63+LV shMIAT group were significantly decreased (P<0. 05), while the proliferation activities and number of angiogenesis in the HUVEC in MG63+LV-MIAT group and IL-4+LV Vector group were significantly increased (P<0. 05). The results of in vivo tumorigenesis experiment of the nude mice showed that compared with LV-Vector group, the volume and weight of transplanted tumor of the mice in LV-MIAT group were significantly increased (P<0. 05), and the positive expression rates of CD163 and CD31 in tumor tissue were significantly increased (P<0. 05)；the volume and mass of transplanted tumor and the positive expression rates of CD163 and CD31 in tumor tissue of the mice in LV-shMIAT group were significantly decreased (P<0. 05). Compared with LV-shMIAT group, the volume and weight of transplanted tumor of the mice in LV-MIAT group were significantly increased (P<0. 05), and the positive expression rates of CD163 and CD31 in tumor tissue were significantly increased (P<0. 05). Conclusion：LncRNA MIAT may regulate the progression of OS by promoting the M2 polarization of macrophages and up-regulating the angiogenesis in tumor tissue. [ABSTRACT FROM AUTHOR]

Objectives: To evaluate the effect of zoledronic acid (ZA) on human umbilical vein endothelial cells (HUVECs) attached to different surfaces. Materials and Methods: A total of three groups were evaluated in this study: sandblasting and acid etching (SLA) + HUVECs; mechanically polished (MP) + HUVECs; and plastic cell culture plates + HUVECs. Scanning electron microscopy, energy‐dispersive X‐ray spectroscopy, surface roughness and water contact angle were tested for titanium surface characterisation. ZA was added at different concentrations (0, 1, 10, 50 and 100 μM). Cell adhesion, proliferation, viability, apoptosis and gene expression were evaluated. Results: Mechanically polished and SLA surfaces showed negative effects on cell adhesion and proliferation and promoted cell apoptosis with 100 μM ZA (p <.05). The highest expression of intercellular adhesion molecule‐1 (ICAM‐1) and angiopoietin‐1 was found on SLA surfaces (p <.01). The lowest expression of platelet‐endothelial cell adhesion molecule‐1 and ICAM‐1 was found on MP surfaces (p <.05). A significant decrease in von Willebrand factor was detected on MP and SLA surfaces (p <.001). Conclusions: Zoledronic acid has an anti‐angiogenic effect on HUVECs attached to titanium implants, while the SLA surface might stimulate HUVECs to express angiogenic and adhesive factor genes despite ZA treatment. [ABSTRACT FROM AUTHOR]

Objectives: To investigate the effects of propranolol on the proliferation, apoptosis, migration, and tube formation ability of human umbilical vein endothelial cells (HUVEC), as well as its impact on the expression of sex-determining region Y-box 18 (SOX18), matrix metalloproteinase-7 (MMP-7), and vascular endothelial growth factor A (VEGFA)., Methods: HUVEC were treated with different concentrations of propranolol, and cell viability was assessed using the CCK-8 method to determine the optimal concentration and treatment duration. The experiment consisted of a control group and groups treated with different concentrations of propranolol (50, 100, 150 μmol/L). Apoptosis, migration, and tube formation of HUVEC were observed using flow cytometry, wound healing assays, and tube formation assays. Western blot and real-time quantitative PCR were used to detect the expression levels of SOX18, MMP-7, and VEGFA proteins and mRNA., Results: Compared to the control group, the apoptosis rate in the propranolol treatment groups increased significantly ( P <0.05), and it rose significantly with increasing drug concentration ( P <0.05). The wound healing rate decreased in the propranolol treatment groups, and both the number of tube formation nodes and total tube length were reduced ( P <0.05). The expression levels of SOX18, MMP-7, and VEGFA proteins and mRNA were downregulated in the propranolol treatment groups ( P <0.05)., Conclusions: Propranolol can inhibit the proliferation, migration, and tube formation ability of HUVEC and promote cell apoptosis, resulting in decreased expression levels of SOX18, MMP-7, and VEGFA.

With the development of magnetic manipulation technology based on magnetic nanoparticles (MNPs), scaffold-free microtissues can be constructed utilizing the magnetic attraction of MNP-labeled cells. The rapid in vitro construction and in vivo vascularization of microtissues with complex hierarchical architectures are of great importance to the viability and function of stem cell microtissues. Endothelial cells are indispensable for the formation of blood vessels and can be used in the prevascularization of engineered tissue constructs. Herein, safe and rapid magnetic labeling of cells was achieved by incubation with MNPs for 1 h, and ultrathick scaffold-free microtissues with different sophisticated architectures were rapidly assembled, layer by layer, in 5 min intervals. The in vivo transplantation results showed that in a stem cell microtissue with trisection architecture, the two separated human umbilical vein endothelial cell (HUVEC) layers would spontaneously extend to the stem cell layers and connect with each other to form a spatial network of functional blood vessels, which anastomosed with the host vasculature. The “hamburger” architecture of stem cell microtissues with separated HUVEC layers could promote vascularization and stem cell survival. This study will contribute to the construction and application of structural and functional tissues or organs in the future.

Dysferlin (DYSF) has drawn much attention due to its involvement in dysferlinopathy and was reported to affect monocyte functions in recent studies. However, the role of DYSF in the pathogenesis of atherosclerotic cardiovascular diseases (ASCVD) and the regulation mechanism of DYSF expression have not been fully studied. In this study, Gene Expression Omnibus (GEO) database and epigenome-wide association study (EWAS) literatures were searched to find the DNA methylation-driven genes (including DYSF) of ASCVD. The hub genes related to DYSF were also identified through weighted correlation network analysis (WGCNA). Regulation of DYSF expression through its promoter methylation status was verified using peripheral blood leucocytes (PBLs) from ASCVD patients and normal controls, and experiments on THP1 cells and Apoe-/- mice. Similarly, the expressions of DYSF related hub genes, mainly contained SELL, STAT3 and TMX1, were also validated. DYSF functions were then evaluated by phagocytosis, transwell and adhesion assays in DYSF knock-down and overexpressed THP1 cells. The results showed that DYSF promoter hypermethylation up-regulated its expression in clinical samples, THP1 cells and Apoe-/- mice, confirming DYSF as a DNA methylation-driven gene. The combination of DYSF expression and methylation status in PBLs had a considerable prediction value for ASCVD. Besides, DYSF could enhance the phagocytosis, migration and adhesion ability of THP1 cells. Among DYSF related hub genes, SELL was proven to be the downstream target of DYSF by wet experiments. In conclusion, DYSF promoter hypermethylation upregulated its expression and promoted monocytes activation, which further participated in the pathogenesis of ASCVD. [ABSTRACT FROM AUTHOR]

In contrast to the stem and fruit of Akebia quinata, A. quinata leaves as a source rich in phenolic compounds with potentially beneficial pharmacological activities have been largely overlooked. To develop and use A. quinata leaves as a resource, we evaluated its potential as a cardiovascular-protective agent. Herein, we investigated the effects and potential mechanisms of A. quinata leaves extract on lipopolysaccharide (LPS)-induced inflammatory responses in human umbilical vein endothelial cells. We found that A. quinata leaves extract pretreatment of 10 μg/mL significantly attenuated LPS-induced protein expression of intercellular adhesion molecule-1, vascular cell adhesion molecule-1. Furthermore, this extract also suppressed LPS-induced phosphorylation of nuclear factor-κB p65. In order to elucidate the chemical profiles of the samples, the HPLC fingerprint was established, and prominent peaks were identified via HPLC–electrospray ionization–mass spectrometry. Multivariate statistical analyses, including hierarchical cluster analysis, principal component analysis, and partial least-squares discriminant analysis, were performed to evaluate the clustering of the samples. It was found that isochlorogenic acid C was a key marker for the classification of A. quinata leaves from the Gongju and Muju city in Korea. Collectively, this study not only suggested the potential of A. quinata leaves as a novel therapeutic candidate for inflammatory cardiovascular disease but also developed a quality control method for A. quinata leaves, which could help to expand the application of A. quinata. [ABSTRACT FROM AUTHOR]

Copyright of Karaelmas Science & Engineering Journal / Karaelmas Fen ve Mühendislik Dergisi is the property of Karaelmas Science & Engineering Journal and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Copyright of Saudi Medical Journal is the property of Saudi Medical Journal and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

This study investigated the release and proteomic profile of tissue factor microparticles (TFMPs) prospectively (up to 6 months) following a myocardial infarction (MI) in a chronic porcine model to establish their utility in tracking cellular level activities that predict physiologic outcomes. Our animal groups (n = 6 to 8 each) consisted of control, noninfarcted (negative control); infarcted only (positive control); and infarcted animals treated with cardiac resynchronization therapy (CRT) and a β-blocker (BB) (metoprolol succinate). The authors found different protein profiles in TFMPs between the control, infarcted only group, and the CRT + BB treated group with predictive impact on the outward phenotype of pathological remodeling after an MI within and between groups. This novel approach of monitoring cellular level activities by profiling the content of TFMPs has the potential of addressing a shortfall of the current crop of cardiac biomarkers, which is the inability to capture composite molecular changes associated with chronic maladaptive signaling in a spatial and temporal manner.

Abstract Background Recent studies have suggested a crucial role of mesenchymal stem cell (MSC)-derived extracellular vesicles (EVs) in ovarian cancer treatment. We, therefore, set out to explore the mechanism through which MSC-derived EVs delivered microRNA-424 (miR-424) to influence the development of ovarian cancer. Methods Bioinformatics analyses were first performed to screen ovarian cancer-related differentially expressed genes and to predict regulatory miRNAs. Then, dual-luciferase reporter gene assay was carried out to verify the relationship between miR-424 and MYB. Subsequently, the characterized MSCs and isolated EVs were co-cultured with ovarian cancer cells, followed by determination of the expression patterns of miR-424, MYB, vascular endothelial growth factor (VEGF), and VEGF receptor (VEGFR), respectively. In addition, the effects of EVs-delivered miR-424 on the proliferation, migration, invasion and tube formation of ovarian cancer cells were assessed using gain- and loss-of-function approaches. Lastly, tumor xenograft was induced in nude mice to illustrate the influence of EVs-loaded miR-424 on ovarian cancer in vivo. Results Our data exhibited that MYB was highly-expressed and miR-424 was poorly-expressed in ovarian cancer. More importantly, MYB was identified as a target gene of miR-424. Additionally, the transfer of miR-424 by MSC-derived EVs was found to repress the proliferation, migration, and invasion of ovarian cancer cells, with a reduction in the expressions of VEGF and VEGFR. Furthermore, MSC-derived EVs over-expressing miR-424 could inhibit the proliferation, migration, and tube formation of human umbilical vein endothelial cells, and also suppressed tumorigenesis and angiogenesis of ovarian tumors in vivo. Conclusion Collectively, our findings indicate that MSC-derived EVs transfer miR-424 to down-regulate MYB, which ultimately led to the inhibition of the tumorigenesis and angiogenesis of ovarian cancer. Hence, this study offers a potential prognostic marker and a therapeutic target for ovarian cancer.

• Capillary structural abnormalities on sun-exposed skin with skin redness unevenness. • Capillary structural abnormalities include dilation and disappearance. • Capillaries are excessively dilated by increased ETBR expression on HUVECs by UVA. • Capillary migration disappears under decreased BMPR2 expression on HUVECs by UVA. Capillary structural abnormalities cause skin disorders. Mottled redness, i.e., skin redness unevenness, may appear on the sun-exposed skin, suggesting capillary structural abnormalities, although its mechanism remains unclear. To observe the capillary structures in the sun-exposed skin where skin redness unevenness is likely to occur, and clarify the mechanism of capillary structural abnormalities. The tissue structures in the skin with skin redness unevenness were observed by LC-OCT. Subsequently, immunostaining of the sun-exposed skin where skin redness unevenness is often observed, was performed. Vascular endothelial cells were UV-irradiated to analyze the expression and functions of genes involved in the capillary structures and morphogenesis. The skin with skin redness unevenness exhibited scattering of dilated tubular tissue and disturbance of distribution uniformity. Immunostaining of the sun-exposed skin that were more likely to be exposed to UV rays also revealed similarly disorder of capillary structures. In addition, UVA-irradiated vascular endothelial cells exhibited increased expression of ETBR, involved in telangiectasia, decreased expression of BMPR2, involved in the morphogenesis and maintenance of the blood vessels, and reduced migration of the capillaries. UV rays alter ETBR and BMPR2 expression in the skin capillaries, and cause partial dilation and decreased migration, resulting in capillary structural abnormalities and causing skin redness unevenness. [ABSTRACT FROM AUTHOR]

Objective To knock out the KNDC1 of human umbilical vein endothelial cells (HUVECs) by CRISPR/Cas9 gene editing technology and observe its anti-aging ability. Methods Five sgRNAs targeting KNDC1 Exon1, Exon2 and Exon3 were designed by online software and were constructed into CRISPR/Cas9 vector. After sequence confirmation, the recombinant plasmids were transfected into HeLa cells and HUVECs. After 48 h, KNDC1 mRNA levels and protein expression levels were detected by real-time PCR and Western blot; cell senescence was measured by SA-β-gal staining; intracellular reactive oxygen species were detected by 2′,7′-dichlorodihydrofluorescein diacetate (DCFH-DA) fluorescent probe. Results The sequencing results showed that all the 5 sgRNAs were inserted into the CRISPR/Cas9 vector, the sequence was correct and these vectors were successfully constructed. After transfecting 5 recombinant vectors into HeLa cells, it was found that the knockout efficiency of vectors No. 4 and No. 5 was higher, and then the recombinant vectors of No. 4 and 5 were transfected into HUVECs. Compared with the control plasmid, the KNDC1 mRNA level and protein expression level in HUVECs transfected with recombinant plasmids 4 and 5 were significantly decreased (P＜0.05). The number of senescent HUVECs and the level of intracellular reactive oxygen species were significantly reduced. Conclusions The KNDC1 of HUVECs can be effectively knocked out by CRISPR/Cas9 technology, and HUVECs with KNDC1 knockout have anti-aging ability.

Abstract Background and aim Inadequate vascularization is a challenge in bone tissue engineering because internal cells are prone to necrosis due to a lack of nutrient supply. Rat bone marrow-derived mesenchymal stem cells (rBMSCs) and human umbilical vein endothelial cells (HUVECs) were cocultured to construct prevascularized bone tissue in osteogenic induction medium (OIM) in vitro. The angiogenic capacity of HUVECs was limited in the coculture system. In this study, the effects of the components in the medium on HUVEC angiogenesis were analyzed. Methods The coculture system was established in OIM. Alizarin red staining and alkaline phosphatase staining were used to assess the osteogenic ability of MSCs. A Matrigel tube assay was used to assess the angiogenic ability of HUVECs in vitro. The proliferation of HUVECs was evaluated by cell counting and CCK-8 assays, and migration was evaluated by the streaked plate assay. The expression levels of angiogenesis-associated genes and proteins in HUVECs were measured by qRT-PCR and Western blotting, respectively. Results Dexamethasone in the OIM suppressed the proliferation and migration of HUVECs, inhibiting the formation of capillary-like structures. Our research showed that dexamethasone stimulated HUVECs to secrete tissue inhibitor of metalloproteinase (TIMP-3), which competed with vascular endothelial growth factor (VEGF-A) to bind to vascular endothelial growth factor receptor 2 (VEGFR2, KDR). This effect was related to inhibiting the phosphorylation of ERK and AKT, which are two downstream targets of KDR. However, under hypoxia, the enhanced expression of hypoxia-inducible factor-1α (HIF-1α) decreased the expression of TIMP-3 and promoted the phosphorylation of KDR, improving HUVEC angiogenesis in the coculture system. Conclusion Coculture of hypoxia-preconditioned HUVECs and MSCs showed robust angiogenesis and osteogenesis in OIM, which has important implications for prevascularization in bone tissue engineering in the future.

One of the earliest events in the development of atherosclerosis is endothelial activation, which is estimated in vitro at the functional level by quantifying monocyte adhesion. This involves the incubation of fluorescently labeled monocytes on top of cultured endothelial cells and quantifying the number of adhered monocytes. Currently, the quantification of adhered monocytes is done using microscopy or by lysing the cells and estimating the fluorescence. Here we present a novel flow cytometry-based method for the quantification of monocyte adhesion. This method could quantify the average number of monocytes adhered to a single endothelial cell after monocyte adhesion assay, and was also sensitive to the level of activation of endothelial cells. Flow cytometry-based quantification requires less time and effort compared with microscopy-based quantification.

Although the contributions of sitagliptin to endothelial dysfunction in diabetes mellitus were previously reported, the mechanisms still undefined. Autophagy plays an important role in the development of diabetes mellitus, but its role in diabetic macrovascular complications is unclear. This study aims to observe the effect of sitagliptin on macrovascular endothelium in diabetes and explore the role of autophagy in this process. Diabetic rats were induced through administration of high-fat diet and intraperitoneal injection of streptozotocin. Then diabetic rats were treated with or without sitagliptin for 12 weeks. Endothelial damage and autophagy were measured. Human umbilical vein endothelial cells were cultured either in normal glucose or in high glucose medium and intervened with different concentrations of sitagliptin. Rapamycin was used to induce autophagy. Cell viability, apoptosis and autophagy were detected. The expressions of proteins in c-Jun N-terminal kinase (JNK)-Bcl-2-Beclin-1 pathway were measured. Sitagliptin attenuated injuries of endothelium in vivo and in vitro. The expression of microtubuleassociated protein 1 light chain 3 II (LC3II) and beclin-1 were increased in aortas of diabetic rats and cells cultured with high-glucose, while sitagliptin inhibited the over-expression of LC3II and beclin-1. In vitro pre-treatment with sitagliptin decreased rapamycin-induced autophagy. However, after pretreatment with rapamycin, the protective effect of sitagliptin on endothelial cells was abolished. Further studies revealed sitagliptin increased the expression of Bcl-2, while inhibited the expression of JNK in vivo. Sitagliptin attenuates injuries of vascular endothelial cells caused by high glucose through inhibiting over-activated autophagy. JNK-Bcl-2-Beclin-1 pathway may be involved in this process. [ABSTRACT FROM AUTHOR]

Maternal diabetes may lead to long-term risks for the offspring. The study aims at identifying the potential crucial genes and pathways associated with foetal metabolism and malformation of gestational diabetes mellitus (GDM). Gene Expression Series 49524 and 87295 were downloaded from Gene Expression Omnibus database, including eight from GDM and eight from non-GDM. A total of 35 differentially expressed genes were identified. Gene ontology functional annotation and signalling pathway analyses were performed. Four hub genes were identified by protein–protein interaction network: SHH, E2F1, STAT1, and HOXA9. The four hub genes were assessed by western blot and real-time quantitative PCR in clinical samples. The results of this data mining and integration help to reveal the pathophysiologic and molecular mechanism imprinted in primary umbilical cord-derived cells from GDM offspring. These genes and pathways identified are potential stratification biomarkers and provide further insight for developing therapeutic intervention for the offspring of diabetic mothers. What is already known on this subject? Maternal diabetes may lead to long-term risks for the offspring. A high glucose environment might change the umbilical cord expression of genes implicated in foetal metabolism and development. However, underlying molecular mechanisms have not been investigated thoroughly. What do the results of this study add? GO functional annotation showed that the biological functions of differentially expressed genes mainly involved in metanephros development, salivary gland morphogenesis, fat cell differentiation, vasculogenesis, muscle cell proliferation, heart morphogenesis and Wnt signalling pathway. Signalling pathway analyses found that these differentially expressed genes mainly implicated in the apoptosis, cell cycle, Hedgehog, P53, and NOTCH signalling pathway. Four hub genes were identified by protein-protein interaction network: SHH, E2F1, STAT1 and HOXA9. What are the implications of these findings for clinical practice and/or further research? The genes and pathways identified in the present study are potential stratification biomarkers and provide further insight for developing therapeutic intervention for the offspring of diabetic mothers. [ABSTRACT FROM AUTHOR]

In 2012, the threshold radiation dose (0.5 Gy) for cardiovascular and cerebrovascular diseases was revised, and this threshold dose may be exceeded during procedures involving radiation such as interventional radiology. Therefore, in addition to regulating radiation dose, it is necessary to develop strategies to prevent and mitigate the development of cardiovascular disease. Cellular senescence is irreversible arrest of cell proliferation. Although cellular senescence is one of the mechanisms for suppressing cancer, it also has adverse effects. For example, senescence of vascular endothelial cells is involved in development of vascular disorders. However, the mechanisms underlying induction of cellular senescence are not fully understood. Therefore, the present study explored the factors involved in the radiation-induced senescence in human umbilical vein endothelial cells (HUVECs). The present study reanalyzed the gene expression data of senescent normal human endothelial cells and fibroblast after irradiation (NCBI Gene Expression Omnibus accession no. GSE130727) and microarray data of HUVECs 24 h after irradiation (NCBI Gene Expression Omnibus accession no. GSE76484). Numerous genes related to viral infection and inflammation were upregulated in radiation-induced senescent cells. In addition, the gene group involved in the retinoic acid-inducible gene-I (RIG-I)-like receptor (RLR) signaling pathway, which plays an important role to induce anti-viral response, was altered in irradiated HUVECs. Therefore, to investigate the involvement of RIG-I and melanoma differentiation-associated gene 5 (MDA5), which are RLRs, in radiation-induced senescence of HUVECs, the protein expression of RIG-I and MDA5 and the activity of senescence-associated β-galactosidase (SA-β-gal), a representative senescence marker, were analyzed. Of note, knockdown of RIG-I in HUVECs significantly decreased radiation-increased proportion of cells with high SA-β-gal activity (i.e., senescent cells), whereas this phenomenon was not observed in MDA5-knockdown cells. Taken together, the present results suggested that RIG-I, but not MDA5, was associated with radiation-induced senescence in HUVECs. [ABSTRACT FROM AUTHOR]

Abstract Sargassum serratifolium ethanolic extract has been known for strong antioxidant and anti-inflammatory properties. We prepared hexane fraction from the ethanolic extract of S. serratifolium (HSS) to improve biological activities. In this study, we investigated the effects of HSS on the inhibition of tumor necrosis factor (TNF)-α-induced monocyte adhesion to human umbilical vein endothelial cells (HUVECs). We found that HSS suppressed the production of cell adhesion molecules such as intracellular adhesion molecule-1 and vascular cell adhesion molecule-1 in TNF-α-induced HUVECs. Moreover, TNF-α-induced production of monocyte chemoattractant protein 1 and keratinocyte chemoattractant was inhibited by HSS treatment. HSS suppressed TNF-α-induced nuclear factor kappa B (NF-κB) activation via preventing proteolytic degradation of inhibitor κB-α. HSS induced the production of heme oxygenase 1 via translocation of Nrf2 into the nucleus in TNF-α-treated HUVECs. Overall, HSS alleviated vascular inflammation through the downregulation of NF-κB activation and the upregulation of Nrf2 activation in TNF-α-induced HUVECs. These results indicate that HSS may be used as therapeutic agents for vascular inflammatory disorders.

In contrast to the stem and fruit of Akebia quinata, A. quinata leaves as a source rich in phenolic compounds with potentially beneficial pharmacological activities have been largely overlooked. To develop and use A. quinata leaves as a resource, we evaluated its potential as a cardiovascular-protective agent. Herein, we investigated the effects and potential mechanisms of A. quinata leaves extract on lipopolysaccharide (LPS)-induced inflammatory responses in human umbilical vein endothelial cells. We found that A. quinata leaves extract pretreatment of 10 μg/mL significantly attenuated LPS-induced protein expression of intercellular adhesion molecule-1, vascular cell adhesion molecule-1. Furthermore, this extract also suppressed LPS-induced phosphorylation of nuclear factor-κB p65. In order to elucidate the chemical profiles of the samples, the HPLC fingerprint was established, and prominent peaks were identified via HPLC–electrospray ionization–mass spectrometry. Multivariate statistical analyses, including hierarchical cluster analysis, principal component analysis, and partial least-squares discriminant analysis, were performed to evaluate the clustering of the samples. It was found that isochlorogenic acid C was a key marker for the classification of A. quinata leaves from the Gongju and Muju city in Korea. Collectively, this study not only suggested the potential of A. quinata leaves as a novel therapeutic candidate for inflammatory cardiovascular disease but also developed a quality control method for A. quinata leaves, which could help to expand the application of A. quinata.

[Human umbilical vascular endothelial cells (HUVECs) underwent ischemia, ischemia/reperfusion and normal control (sham) treatment and marked as group I, IR and NC, respectively, were detected by quantitative proteomics and bioinformatics analyses. Proteins in Beclin-1/LC3-II-dependent canonical macroautophagy pathway were verified in details. The significantly upregulated proteins encoded by autophagy-related genes (ATGs) included ATG2A, ATG3, ATG4B, ATG5, ATG7, ATG9A, ATG12, ATG16 and ATG101. The significantly enhanced lysosomal proteins comprised Cathepsin B, Cathepsin D, lysosome-associated membrane protein 1 (LAMP1) and LAMP2. However, the differentially expressed proteins excluded Beclin-1, microtubule-associated protein light chain 3 (LC3)-I and LC3-II. Western blot analyses verified that the protein expressions of Beclin-1, LC3-I and LC3-II were neither upregulated nor downregulated in ischemia-challenged HUVECs.]

In a randomized trial, higher all-cause and cardiovascular mortality was observed in treatment with febuxostat than with allopurinol in patients with coexisting gout and serious cardiovascular conditions. In this study, we focus on an intervention of febuxostat or allopurinol as an anti-inflammatory treatment to reduce the transcription of nuclear factor-kappa B (NF-κB) and production of relevant inflammatory factors. We evaluated the effect of febuxostat on vascular cell adhesion protein 1 (VCAM-1) induction in cultured human umbilical vein endothelial cells (HUVECs). Cells were exposed to tumor necrosis factor (TNF)-α (10 ng/mL) treatment for 24 h. Febuxostat or allopurinol (0.1–100 μM) was added to the bath medium 15 min before TNF-α treatment. VCAM-1 levels in HUVECs increased after 24-h TNF-α treatment (n = 4). Febuxostat and allopurinol significantly suppressed VCAM-1 induced by treatment with TNF-α in a dose-dependent manner (p < 0.05, n = 4). Furthermore, these drugs suppressed the NF-κB protein levels in the nucleus 4 h after TNF-α treatment (n = 3 or 4). Our results suggest that TNF-α induces VCAM-1 production via NF-κB, which can be blocked by febuxostat or allopurinol. The effect of febuxostat treatment on cardiovascular events may be associated with protection against the infiltration of lymphocytes or monocytes through VCAM-1 induction in inflamed endothelial cells such as arterial sclerosis. [ABSTRACT FROM AUTHOR]

Background: Recent studies have suggested a crucial role of mesenchymal stem cell (MSC)-derived extracellular vesicles (EVs) in ovarian cancer treatment. We, therefore, set out to explore the mechanism through which MSC-derived EVs delivered microRNA-424 (miR-424) to influence the development of ovarian cancer.Methods: Bioinformatics analyses were first performed to screen ovarian cancer-related differentially expressed genes and to predict regulatory miRNAs. Then, dual-luciferase reporter gene assay was carried out to verify the relationship between miR-424 and MYB. Subsequently, the characterized MSCs and isolated EVs were co-cultured with ovarian cancer cells, followed by determination of the expression patterns of miR-424, MYB, vascular endothelial growth factor (VEGF), and VEGF receptor (VEGFR), respectively. In addition, the effects of EVs-delivered miR-424 on the proliferation, migration, invasion and tube formation of ovarian cancer cells were assessed using gain- and loss-of-function approaches. Lastly, tumor xenograft was induced in nude mice to illustrate the influence of EVs-loaded miR-424 on ovarian cancer in vivo.Results: Our data exhibited that MYB was highly-expressed and miR-424 was poorly-expressed in ovarian cancer. More importantly, MYB was identified as a target gene of miR-424. Additionally, the transfer of miR-424 by MSC-derived EVs was found to repress the proliferation, migration, and invasion of ovarian cancer cells, with a reduction in the expressions of VEGF and VEGFR. Furthermore, MSC-derived EVs over-expressing miR-424 could inhibit the proliferation, migration, and tube formation of human umbilical vein endothelial cells, and also suppressed tumorigenesis and angiogenesis of ovarian tumors in vivo.Conclusion: Collectively, our findings indicate that MSC-derived EVs transfer miR-424 to down-regulate MYB, which ultimately led to the inhibition of the tumorigenesis and angiogenesis of ovarian cancer. Hence, this study offers a potential prognostic marker and a therapeutic target for ovarian cancer. [ABSTRACT FROM AUTHOR]

Background/Aims: Increasing evidence indicates that microRNAs (miRNAs) play important roles in Kawasaki disease (KD). Our previous study demonstrated that hsa-miR-27b-3p (miR-27b) was up-regulated in KD serum. However, the specific role of miR-27b in KD remains unclear. We aimed to investigate that miR-27b could be a biomarker and therapeutic target for KD treatment. As well, the specific mechanism of miR-27b effecting endothelial cell functions was studied. Methods: The expression of miR-27b and Smad7 was measured by qRT-PCR. Gain-of-function strategy was used to observe the effect of miR-27b on human umbilical vein endothelial cells (HUVECs) proliferation and migration. Bioinformatics analyses were applied to predict miR-27b targets and then we verified Smad7 by a luciferase reporter assay. Western blot was performed to detect the protein expression of Smad7, PCNA, MMP9, MMP12 and TGF-β-related genes. Results: We confirmed that miR-27b was shown to be dramatically up-regulated in KD serum and KD serum-treated HUVECs and that elevated expression of miR-27b suppressed the proliferation and migration of HUVECs. Furthermore, our results verified that miR-27b mediated cell functions by affecting the TGF-β via targeting Smad7 in HUVECs. Conclusion: These results suggested that up-regulated miR-27b had a protective role in HUVECs proliferation and migration via targeting Smad7 and affecting TGF-β pathway. Therefore, miR-27b represented a potential biomarker for KD and may serve as a promising therapeutic target for KD treatment.

Angiogenesis is the formation of new capillaries from pre-existing blood vessels and participates in proper vasculature development. In pathological conditions such as cancer, abnormal angiogenesis takes place. Angiogenesis is primarily carried out by endothelial cells, the innermost layer of blood vessels. The vascular endothelial growth factor-A (VEGF-A) and its receptor-2 (VEGFR-2) trigger most of the mechanisms activating and regulating angiogenesis, and have been the targets for the development of drugs. However, most experimental assays assessing angiogenesis rely on animal models. We report an in vitro model using a microvessel-on-a-chip. It mimics an effective endothelial sprouting angiogenesis event triggered from an initial microvessel using a single angiogenic factor, VEGF-A. The angiogenic sprouting in this model is depends on the Notch signaling, as observed in vivo. This model enables the study of anti-angiogenic drugs which target a specific factor/receptor pathway, as demonstrated by the use of the clinically approved sorafenib and sunitinib for targeting the VEGF-A/VEGFR-2 pathway. Furthermore, this model allows testing simultaneously angiogenesis and permeability. It demonstrates that sorafenib impairs the endothelial barrier function, while sunitinib does not. Such in vitro human model provides a significant complimentary approach to animal models for the development of effective therapies.

Wound healing is a complex physiological process in which fibrocytes serve a vital role. However, the mechanism underlying the recruitment of fibrocytes during wound healing remains largely unknown. The present study aimed to investigate whether endothelial cells are involved in the recruitment of fibrocytes in wound healing. To mimic the in vivo angiogenic process, a co‑culture system consisting of endothelial cells and fibrocytes was achieved using a permeable Transwell co‑culture system. The expression of chemokines produced by endothelial cells with or without co‑culture was then measured using a gene chip. Based on the dataset from chip analysis, chemokine ligand 15 (CCL15) produced by endothelial cells was identified, which likely serves a regulatory role in mediating the transmigration of fibrocytes. Overexpression of CCL15 in endothelial cells or chemokine receptor 1 (CCR1) in fibrocytes promoted the transmigration of fibrocytes, whilst silencing the expression of CCL15 in endothelial cells or that of CCR1 in fibrocytes attenuated the transmigration of fibrocytes. Results from the present study suggested that the CCL15‑CCR1 axis between endothelial cells and fibrocytes serves a vital role in mediating the recruitment of fibrocytes during wound healing. [ABSTRACT FROM AUTHOR]

Background: Cardiovascular disease (CVD) is one of the major cause of mortality in diabetic patients. Evidence suggests that hyperglycemia in diabetic patients contributes to increased risk of CVD. This study is to investigate the therapeutic effects of melatonin on glucose-treated human umbilical vein endothelial cells (HUVEC) and provide insights on the underlying mechanisms. Materials and methods: Cell viability was determined using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Reactive oxygen species (ROS) and membrane potential was detected using 2′,7′-dichlorofluorescein diacetate and 5,5′,6,6′-tetrachloro-1,1′,3,3′-tetraethylbenzimidazolcarbocyanine iodide (JC-1) dye staining, respectively. While, cell apoptosis was determined by Annexin-V staining and protein expression was measured using Western blot. Results: Our results suggested that melatonin inhibited glucose-induced ROS elevation, mitochondria dysfunction and apoptosis on HUVEC. Melatonin inhibited glucose-induced HUVEC apoptosis via PI3K/Akt signaling pathway. Activation of Akt further activated BcL-2 pathway through upregulation of Mcl-1 expression and downregulation Bax expression in order to inhibit glucose-induced HUVEC apoptosis. Besides that, melatonin promoted downregulation of oxLDL/LOX-1 in order to inhibit glucose-induced HUVEC apoptosis. Conclusions: In conclusion, our results suggested that melatonin exerted vasculoprotective effects against glucose-induced apoptosis in HUVEC through PI3K/Akt, Bcl-2 and oxLDL/LOX-1 signaling pathways. [ABSTRACT FROM AUTHOR]

Background and aim: Inadequate vascularization is a challenge in bone tissue engineering because internal cells are prone to necrosis due to a lack of nutrient supply. Rat bone marrow-derived mesenchymal stem cells (rBMSCs) and human umbilical vein endothelial cells (HUVECs) were cocultured to construct prevascularized bone tissue in osteogenic induction medium (OIM) in vitro. The angiogenic capacity of HUVECs was limited in the coculture system. In this study, the effects of the components in the medium on HUVEC angiogenesis were analyzed. Methods: The coculture system was established in OIM. Alizarin red staining and alkaline phosphatase staining were used to assess the osteogenic ability of MSCs. A Matrigel tube assay was used to assess the angiogenic ability of HUVECs in vitro. The proliferation of HUVECs was evaluated by cell counting and CCK-8 assays, and migration was evaluated by the streaked plate assay. The expression levels of angiogenesis-associated genes and proteins in HUVECs were measured by qRT-PCR and Western blotting, respectively. Results: Dexamethasone in the OIM suppressed the proliferation and migration of HUVECs, inhibiting the formation of capillary-like structures. Our research showed that dexamethasone stimulated HUVECs to secrete tissue inhibitor of metalloproteinase (TIMP-3), which competed with vascular endothelial growth factor (VEGF-A) to bind to vascular endothelial growth factor receptor 2 (VEGFR2, KDR). This effect was related to inhibiting the phosphorylation of ERK and AKT, which are two downstream targets of KDR. However, under hypoxia, the enhanced expression of hypoxia-inducible factor-1α (HIF-1α) decreased the expression of TIMP-3 and promoted the phosphorylation of KDR, improving HUVEC angiogenesis in the coculture system. Conclusion: Coculture of hypoxia-preconditioned HUVECs and MSCs showed robust angiogenesis and osteogenesis in OIM, which has important implications for prevascularization in bone tissue engineering in the future. [ABSTRACT FROM AUTHOR]

Lack of bone grafts appeals for bone augmentation solutions. We aimed at osteogenic differentiation of human adipose stem cells (hASCs) and microvascularization in coculture with human umbilical vein endothelial cells (HUVECs) embedded in three‐dimensional (3D) gellan gum (GG) and collagen type I (COL) hydrogel mixture. We compared endothelial growth medium‐2 (EGM‐2) and bioactive glass extract‐based endothelial and osteogenic medium (BaG EM‐OM) for vascularized bone‐like graft development in vitro. Cell viability, cell number, and osteogenic and endothelial gene expression were analyzed. Mineralized hydroxyapatite residues, immunocytochemical staining of endothelial marker CD31 production and late osteogenic marker osteocalcin were imaged. With both media, good cell viability was observed within 3D hydrogel. EGM‐2 condition induced significantly higher cell number compared to BaG EM‐OM condition at both 7 and 14 days. Interestingly, both media supported osteogenic as well as endothelial marker gene expression. Moreover, formation of reticulated cellular structures was observed in both EGM‐2 and BaG EM‐OM conditions. However, hydroxyapatite mineralization and strong osteocalcin staining were detected only in BaG EM‐OM condition. Importantly, strong production of CD31 and elongated tube‐like structures were apparent in EGM‐2 culture alone. In conclusion, we demonstrated efficient hASC osteogenic differentiation and microvessel‐like network formation in coculture with HUVECs. [ABSTRACT FROM AUTHOR]