Migration Of Human Endothelial Cells Research Articles

Biomimetic tri-layered small-diameter vascular grafts with decellularized extracellular matrix promoting vascular regeneration and inhibiting thrombosis with the salidroside.

Small-diameter vascular grafts (SDVGs) are urgently required for clinical applications. Constructing vascular grafts mimicking the defining features of native arteries is a promising strategy. Here, we constructed a tri-layered vascular graft with a native artery decellularized extracellular matrix (dECM) mimicking the component of arteries. The porcine thoracic aorta was decellularized and milled into dECM powders from the differential layers. The intima and media dECM powders were blended with poly (L-lactide-co-caprolactone) (PLCL) as the inner and middle layers of electrospun vascular grafts, respectively. Pure PLCL was electrospun as a strengthening sheath for the outer layer. Salidroside was loaded into the inner layer of vascular grafts to inhibit thrombus formation. In vitro studies demonstrated that dECM provided a bioactive milieu for human umbilical vein endothelial cell (HUVEC) extension adhesion, proliferation, migration, and tube-forming. The in vivo studies showed that the addition of dECM could promote endothelialization, smooth muscle regeneration, and extracellular matrix deposition. The salidroside could inhibit thrombosis. Our study mimicked the component of the native artery and combined it with the advantages of synthetic polymer and dECM which provided a promising strategy for the design and construction of SDVGs.

The marriage of immunomodulatory, angiogenic, and osteogenic capabilities in a piezoelectric hydrogel tissue engineering scaffold for military medicine

BackgroundMost bone-related injuries to grassroots troops are caused by training or accidental injuries. To establish preventive measures to reduce all kinds of trauma and improve the combat effectiveness of grassroots troops, it is imperative to develop new strategies and scaffolds to promote bone regeneration.MethodsIn this study, a porous piezoelectric hydrogel bone scaffold was fabricated by incorporating polydopamine (PDA)-modified ceramic hydroxyapatite (PDA-hydroxyapatite, PHA) and PDA-modified barium titanate (PDA-BaTiO3, PBT) nanoparticles into a chitosan/gelatin (Cs/Gel) matrix. The physical and chemical properties of the Cs/Gel/PHA scaffold with 0–10 wt% PBT were analyzed. Cell and animal experiments were performed to characterize the immunomodulatory, angiogenic, and osteogenic capabilities of the piezoelectric hydrogel scaffold in vitro and in vivo.ResultsThe incorporation of BaTiO3 into the scaffold improved its mechanical properties and increased self-generated electricity. Due to their endogenous piezoelectric stimulation and bioactive constituents, the as-prepared Cs/Gel/PHA/PBT hydrogels exhibited cytocompatibility as well as immunomodulatory, angiogenic, and osteogenic capabilities; they not only effectively induced macrophage polarization to M2 phenotype but also promoted the migration, tube formation, and angiogenic differentiation of human umbilical vein endothelial cells (HUVECs) and facilitated the migration, osteo-differentiation, and extracellular matrix (ECM) mineralization of MC3T3-E1 cells. The in vivo evaluations showed that these piezoelectric hydrogels with versatile capabilities significantly facilitated new bone formation in a rat large-sized cranial injury model. The underlying molecular mechanism can be partly attributed to the immunomodulation of the Cs/Gel/PHA/PBT hydrogels as shown via transcriptome sequencing analysis, and the PI3K/Akt signaling axis plays an important role in regulating macrophage M2 polarization.ConclusionThe piezoelectric Cs/Gel/PHA/PBT hydrogels developed here with favorable immunomodulation, angiogenesis, and osteogenesis functions may be used as a substitute in periosteum injuries, thereby offering the novel strategy of applying piezoelectric stimulation in bone tissue engineering for the enhancement of combat effectiveness in grassroots troops.

SALL4 promotes angiogenesis in gastric cancer by regulating VEGF expression and targeting SALL4/VEGF pathway inhibits cancer progression

BackgroundSpalt-like protein 4 (SALL4) is a stemness-related transcription factor whose abnormal re-expression contributes to cancer initiation and progression. However, the role of SALL4 in cancer angiogenesis remains unknown.MethodsAnalyses of clinical specimens via TCGA datasets were performed to determine the expression level and clinical significance of SALL4 in STAD (Stomach Adenocarcinoma). SALL4 knockdown, knockout, and overexpression were achieved by siRNA, CRISPR/Cas9, and plasmid transfection. The effects of conditioned medium (CM) from SALL4 knockdown or overexpression of gastric cancer cells on endothelial cell proliferation, migration, and tube formation were investigated by CCK-8 assay, transwell migration assay, and tube formation assay. The regulation of VEGF gene expression by SALL4 was studied by qRT-PCR, western blot, chromatin immunoprecipitation (ChIP) assay, and electrophoretic mobility shift assay (EMSA). Engineered exosomes from 293T cells loaded with si-SALL4-B and thalidomide were produced to test their therapeutic effect on gastric cancer progression.ResultsSALL4 expression was increased in STAD and positively correlated with tumor progression and poor prognosis. SALL4-B knockdown or knockout decreased while over-expression increased the promotion of human umbilical vein endothelial cells (HUVEC) cell proliferation, migration, and tube formation by gastric cancer cell-derived CM. Further investigation revealed a widespread association of SALL4 with angiogenic gene transcription through the TCGA datasets. Additionally, SALL4-B knockdown reduced, while over-expression enhanced the expression levels of VEGF-A, B, and C genes. The results of ChIP and EMSA assays indicated that SALL4 could directly bind to the promoters of VEGF-A, B, and C genes and activate their transcription, which may be associated with increased histone H3-K79 and H3-K4 modifications in their promoter regions. Furthermore, si-SALL4-B and thalidomide-loaded exosomes could be efficiently uptaken by gastric cancer cells and significantly reduced SALL4-B and Vascular Endothelial Growth Factor (VEGF) expression levels in gastric cancer cells, thus inhibiting the pro-angiogenic role of their derived CM.ConclusionThese findings suggest that SALL4 plays an important role in angiogenesis by transcriptionally regulating VEGF expression. Co-delivery of the functional siRNA and anticancer drug via exosomes represents a useful approach to inhibiting cancer angiogenesis by targeting SALL4/VEGF pathway.

Disintegrin-like Protein Strategy to Inhibit Aggressive Triple-Negative Breast Cancer.

Venoms are a rich source of bioactive compounds, and among them is leberagin-C (Leb-C), a disintegrin-like protein derived from the venom of Macrovipera lebetina transmediterrannea snakes. Leb-C has shown promising inhibitory effects on platelet aggregation. Previous studies have demonstrated that this SECD protein specifically targets α5β1, αvβ3, and αvβ6 integrins through a mimic mechanism of RGD disintegrins. In our current study, we focused on exploring the potential effects of Leb-C on metastatic breast cancer. Our findings revealed that Leb-C disrupted the adhesion, migration, and invasion capabilities of MDA-MB-231 breast cancer cells and its highly metastatic D3H2LN sub-population. Additionally, we observed significant suppression of adhesion, migration, and invasion of human umbilical vein endothelial cells (HUVECs). Furthermore, Leb-C demonstrated a strong inhibitory effect on fibroblast-growth-factor-2-induced proliferation of HUVEC. We conducted in vivo experiments using nude mice and found that treatment with 2 µM of Leb-C resulted in a remarkable 73% reduction in D3H2LN xenograft tumor size. Additionally, quantification of intratumor microvessels revealed a 50% reduction in tumor angiogenesis in xenograft after 21 days of twice-weekly treatment with 2 µM of Leb-C. Collectively, these findings suggest the potential utility of this disintegrin-like protein for inhibiting aggressive and resistant metastatic breast cancer.

Thermosensitive hydrogel coupled with sodium ascorbyl phosphate promotes human umbilical cord-derived mesenchymal stem cell-mediated skin wound healing in mice.

Poor survival and restricted function of transplanted stem cells are regarded as limiting their efficacy in wound recovery greatly. Consequently, it is necessary to identify innovative therapeutic strategies to solve these issues. Firstly, the biological effect of PF-127 hydrogel alone and in combination with SAP on the survival, and migration of cultured HUCMSCs was assessed by cell viability, apoptosis, and scratch wound assays. S. aureus and E. coli were used to evaluate the antibacterial activity of PF-127 plus SAP combination. Further, the ability of HUCMSCs-conditioned medium (HUCMSCs-CM) to promote the angiogenesis and migration of human umbilical vein endothelial cells (HUVECs) in vitro was evaluated using tube formation and transwell migration assays. Finally, the HUCMSCs embedded in PF-127 plus SAP scaffold were administered onto mice's excisional cutaneous wound bed. Histological and immunohistochemical analyses were employed to investigate the wound healing capacity as well as cellular responses of PF-127/HUCMSCs/SAP hydrogel. PF-127 showed cytotoxicity on HUCMSCs, whereas the addition of SAP significantly promoted cell viability and alleviated apoptosis of HUCMSCs encapsulated in PF-127 hydrogel in vitro. SAP supplementation substantially abrogated the inhibiting effect of PF-127 on the migration of HUCMSCs in vitro. The combination of PF-127 and SAP exerted an obvious bacteriostatic function on S. aureus and E. coli. Moreover, the co-treatment with SAP could remarkably enhance the stimulative effect of HUCMSCs-CM on the angiogenesis and migration of HUVECs in vitro. PF-127 combined SAP-embedded HUCMSCs transplantation resulted in a potently accelerated wound healing process, promoted the number of proliferating cells and newly formed blood vessels, as well as enhanced expression of vascular endothelial growth factor. PF-127 coupled with SAP contributes to HUCMSCs-mediated traumatic wound closure in mice by promoting cell survival, antibacterial action, and angiogenesis. Our results offered a theoretical foundation for the clinical treatment of traumatic skin defects.

HAND2-AS1 associates with outcomes of acute coronary syndrome and regulates cell viability of vascular endothelial cells.

Acute coronary syndrome (ACS) is an emergency and severe disorder of the cardiovascular system. This paper assessed the expression of plasma HAND2-AS1 in patients with ACS, researched its diagnostic and prognostic significance, and studied its possible mechanism for participating in ACS. The concentration of HAND2-AS1 of 101 included patients with ACS was certified by qRT-PCR and its possible diagnostic function was revealed by the receiver operating characteristic (ROC) curve. All patients were followed up for 6months after percutaneous coronary intervention (PCI) therapy and Kaplan-Meier (K-M) curve and COX regression analysis was performed to estimate the short-term prognostic value of HAND2-AS1 in ACS. The interrelationship between HAND2-AS1 and Gensini score and endothelial injury was identified via Pearson correlation. The function of HAND2-AS1 on the viability, migration, and apoptosis of human umbilical vein endothelial cells (HUVECs) was estimated by the Cell Counting Kit-8 (CCK-8), Transwell chamber, and flow cytometry. In ACS patients, the expression of serum HAND2-AS1 was prominently decreased and closely correlated with the Gensini score. The decreased HAND2-AS1 expression was of diagnostic significance. Declined plasma HAND2-AS1 was observed in patients with the major adverse cardio-cerebrovascular event (MACCE) and was an independent risk for the poor prognosis of ACS patients. In the cell model, upregulation of HAND2-AS1 improved cell viability and migration and inhibited cell apoptosis. HAND2-AS1 was an independent biomarker for the diagnosis and prognosis of ACS. HAND2-AS1 might be involved in ACS development by regulating endothelial damage.

Metabolomic Profiling of Long-Chain Polyunsaturated Fatty Acid Oxidation in Adults with Retinal Vein Occlusion: A Case-Control Study

BackgroundLong-chain polyunsaturated fatty acids (LCPUFAs) and their metabolites are closely related to neovascular eye diseases. However, the clinical significance of their oxylipins in retinal vein occlusion (RVO) remains inconclusive. ObjectivesThis case-control study aimed to explore metabolomic profiles of LCPUFA oxidation in RVO and to identify potential indicators for diagnosis and pathologic progression. MethodsThe plasma concentrations of ω-3 (n–3) and ω-6 (n–6) LCPUFA and their oxylipins in 44 adults with RVO and 36 normal controls were analyzed using ultraperformance liquid chromatography tandem mass spectrometry. Univariate analysis combined with principal component and orthogonal projections to latent structure discriminant analysis was used to screen differential metabolites. Aortic ring and choroidal explant sprouting assays were used to investigate the effects of 5-oxo-eicosatetraenoic acids (ETE) on angiogenesis ex vivo. Tubule formation and wound healing assays were performed to verify its effects on human retinal microvascular endothelial cell functions. ResultsHigher ω-6 and lower ω-3 LCPUFA plasma concentrations were measured in the adults with RVO compared with control (odds ratio [OR]: 2.34; 95% confidence interval [CI]: 1.42, 3.86; P < 0.001; OR: 0.28; 95% CI: 0.15, 0.51; P < 0.001). Metabolomic analysis revealed 20 LCPUFA and their oxylipins dysregulated in RVO, including increased arachidonic acid (ω-6, OR: 1.85; 95% CI: 1.18, 2.90; P < 0.001) and its lipoxygenase product 5-oxo-ETE (OR: 11.76; 95% CI: 3.73, 37.11; P < 0.001), as well as decreased docosahexaenoic acid (ω-3, OR: 0.13; 95% CI: 0.05, 0.33; P < 0.001). Interestingly, 5-oxo-ETE was downregulated in ischemic compared with nonischemic central RVO. Exogenous 5-oxo-ETE attenuated aortic ring and choroidal explant sprouting and inhibited tubule formation and migration of human retinal microvascular endothelial cells in a dose-dependent manner, possibly via suppressing the vascular endothelial growth factor signaling pathway. ConclusionsThe plasma concentrations of ω-6 and ω-3 LCPUFA and their oxylipins were associated with RVO. The ω-6 LCPUFA-derived metabolite 5-oxo-ETE was a potential marker of RVO development and progression.

Human Adipose-Derived Mesenchymal Stem Cell-Secreted Extracellular Matrix Coating on a Woven Nanotextile Vascular Patch for Improved Endothelial Cell Response.

Biomedical implants possessing the structural and functional characteristics of extracellular matrix (ECM) are pivotal for vascular applications. This study investigated the potential of recreating a natural ECM-like structural and functional environment on the surface of biodegradable polymeric nanotextiles for vascular implants. Human adipose-derived mesenchymal stem cells (MSCs) were grown on a suitably engineered polycaprolactone (PCL) nanofibrous textile and were allowed to modify its surface through the deposition of MSC-specific ECM. This surface-modified nanotextile showed mechanical characteristics and functionality appropriate for vascular patch material. The uniformity of ECM coating significantly improved the viability, proliferation, and migration of human endothelial cells compared to bare and xenogeneic collagen-coated PCL nanotextile patches. Thus, a polymeric nanotextile, which is surface modified using MSC-driven ECM, provided a rapid and improved endothelialization, thereby suggesting its potential for vascular patch applications.

Inhibition of Monoacylglycerol Lipase Decreases Angiogenic Features of Endothelial Cells via Release of Tissue Inhibitor of Metalloproteinase-1 from Lung Cancer Cells.

Despite the well-described anticarcinogenic effects of endocannabinoids, the influence of the endocannabinoid system on tumor angiogenesis is still debated. In the present study, conditioned medium (CM) from A549 and H358 lung cancer cells treated with ascending concentrations of the monoacylglycerol lipase (MAGL) inhibitor JZL184 and 2-arachidonoylglycerol (2-AG), a prominent MAGL substrate, caused a concentration-dependent reduction in human umbilical vein endothelial cell (HUVEC) migration and tube formation compared with CM from vehicle-treated cancer cells. Comparative experiments with MAGL inhibitors JW651 and MJN110 showed the same results. On the other hand, the angiogenic properties of HUVECs were not significantly altered by direct stimulation with JZL184 or 2-AG or by exposure to CM of JZL184- or 2-AG-treated non-cancerous bronchial epithelial cells (BEAS-2B). Inhibition of HUVEC migration and tube formation by CM of JZL184- and 2-AG-treated A549 cells was abolished in the presence of the CB1 antagonist AM-251. Increased release of tissue inhibitor of metalloproteinase-1 (TIMP-1) from JZL184- or 2-AG-stimulated A549 or H358 cells was shown to exert an antiangiogenic effect on HUVECs, as confirmed by siRNA experiments. In addition, JZL184 caused a dose-dependent regression of A549 tumor xenografts in athymic nude mice, which was associated with a decreased number of CD31-positive cells and upregulation of TIMP-1-positive cells in xenograft tissue. In conclusion, our data suggest that elevation of 2-AG by MAGL inhibition leads to increased release of TIMP-1 from lung cancer cells, which mediates an antiangiogenic effect on endothelial cells.

The endothelial-to-mesenchymal transition changes the focal adhesion site proteins levels and the SLRP-lumican level in HMEC-1 cell line

Scleroderma, the chronic autoimmune disease is a consequence of inflammation in the connective tissue. Prolonged duration affects formation of compact connective tissue strands (scarring) within the target organ. Endothelial cells undergoing endothelial-to-mesenchymal transition (EndMT) are the source of fibroblast phenotype-resembling cells. EndMT contributes to reorganization of the focal adhesion proteins (FA), including integrins, and intensive extracellular matrix (ECM) remodelling. However, in endothelial cells, the relationship between EndMT and the interaction of integrin receptors with lumican - a component of ECM, is still unclear. Our findings indicate that at the early stages of EndMT caused by Snail-1 transcription factor overexpression, the level of the β1 integrin subunit and its phosphorylation are elevated. Simultaneously, the changes in the level of proteins that build FAs and promote activation of integrin receptors as well as a decrease in lumican quantity were observed. These modulations contributed to increased migration of human microvascular endothelial cells, HMEC-1. Our findings were achieved by WB, ELISA and wound healing assay. Taken altogether, transfection of HMEC-1 cells with Snail-1 plasmids inducing the early stages of EndMT results in the increase of total FAK and integrin β1 phosphorylation as well as cell migration: phenomena which are modulated by interaction with lumican.

Curcumin-loaded porous scaffold: an anti-angiogenic approach to inhibit endochondral ossification

Bone marrow stem cells (BMSCs) are recognized for their robust proliferative capabilities and multidirectional differentiation potential. Ectopic endochondral ossification of BMSC-generated cartilage in subcutaneous environments is a concern associated with vascularization. Hence, devising a reliable strategy to inhibit vascularization is crucial. In this study, an anti-angiogenic drug, curcumin (Cur), was encapsulated into gelatin to create a porous Cur/Gelatin scaffold, with the aim of inhibiting vascular invasion and preventing endochondral ossification of BMSC-regenerated cartilage. In vitro wound healing tests demonstrated that a 30 μM Cur solution could inhibit the migration and growth of human umbilical vein endothelial cells without impeding BMSCs migration and growth. Compared to the gelatin scaffold, our findings verified that the Cur/Gelatin scaffold significantly inhibited vascular invasion after being subcutaneously implanted into rabbits for 12 weeks, as evidenced by gross observation and immunofluorescence CD31 staining. Moreover, both the porous gelatin and Cur/Gelatin scaffolds were populated with BMSCs and underwent in vitro chondrogenic cultivation to produce cartilage, followed by subcutaneous implantation in rabbits for 12 weeks. Histological examinations (including HE, Safranin-O/Fast Green, toluidine blue, and immunohistochemical COL II staining) revealed that the BMSC-generated cartilage in the gelatin group exhibited prominent endochondral ossification. In contrast, the BMSC-generated cartilage in the Cur/Gelatin group maintained cartilage features, such as cartilage matrix and lacunar structure. This study suggests that Cur-loaded scaffolds offer a reliable platform to inhibit endochondral ossification of BMSC-generated cartilage.

Computer simulation approach to the identification of visfatin-derived angiogenic peptides.

Angiogenesis plays an essential role in various normal physiological processes, such as embryogenesis, tissue repair, and skin regeneration. Visfatin is a 52 kDa adipokine secreted by various tissues including adipocytes. It stimulates the expression of vascular endothelial growth factor (VEGF) and promotes angiogenesis. However, there are several issues in developing full-length visfatin as a therapeutic drug due to its high molecular weight. Therefore, the purpose of this study was to develop peptides, based on the active site of visfatin, with similar or superior angiogenic activity using computer simulation techniques.Initially, the active site domain (residues 181∼390) of visfatin was first truncated into small peptides using the overlapping technique. Subsequently, the 114 truncated small peptides were then subjected to molecular docking analysis using two docking programs (HADDOCK and GalaxyPepDock) to generate small peptides with the highest affinity for visfatin. Furthermore, molecular dynamics simulations (MD) were conducted to investigate the stability of the protein-ligand complexes by computing root mean square deviation (RSMD) and root mean square fluctuation(RMSF) plots for the visfatin-peptide complexes. Finally, peptides with the highest affinity were examined for angiogenic activities, such as cell migration, invasion, and tubule formation in human umbilical vein endothelial cells (HUVECs). Through the docking analysis of the 114 truncated peptides, we screened nine peptides with a high affinity for visfatin. Of these, we discovered two peptides (peptide-1: LEYKLHDFGY and peptide-2: EYKLHDFGYRGV) with the highest affinity for visfatin. In an in vitrostudy, these two peptides showed superior angiogenic activity compared to visfatin itself and stimulated mRNA expressions of visfatin and VEGF-A. These results show that the peptides generated by the protein-peptide docking simulation have a more efficient angiogenic activity than the original visfatin.

Methylglyoxal Impairs the Pro-Angiogenic Ability of Mouse Adipose-Derived Stem Cells (mADSCs) via a Senescence-Associated Mechanism.

Adipose-derived stem cells (ADSCs) play a crucial role in angiogenesis and repair of damaged tissues. However, in pathological conditions including diabetes, ADSC function is compromised. This work aims at evaluating the effect of Methylglyoxal (MGO), a product of chronic hyperglycemia, on mouse ADSCs' (mADSCs) pro-angiogenic function and the molecular mediators involved. The mADSCs were isolated from C57bl6 mice. MGO-adducts and p-p38 MAPK protein levels were evaluated by Western Blot. Human retinal endothelial cell (hREC) migration was analyzed by transwell assays. Gene expression was measured by qRT-PCR, and SA-βGal activity by cytofluorimetry. Soluble factor release was evaluated by multiplex assay. MGO treatment does not impair mADSC viability and induces MGO-adduct accumulation. hREC migration is reduced in response to both MGO-treated mADSCs and conditioned media from MGO-treated mADSCs, compared to untreated cells. This is associated with an increase of SA-βGal activity, SASP factor release and p53 and p21 expression, together with a VEGF- and PDGF-reduced release from MGO-treated mADSCs and a reduced p38-MAPK activation in hRECs. The MGO-induced impairment of mADSC function is reverted by senolytics. In conclusion, MGO impairs mADSCs' pro-angiogenic function through the induction of a senescent phenotype, associated with the reduced secretion of growth factors crucial for hREC migration.

ABCB5+ Limbal Epithelial Stem Cells Inhibit Developmental but Promote Inflammatory (Lymph) Angiogenesis While Preventing Corneal Inflammation.

The limbus, the vascularized junction between the cornea and conjunctiva, is thought to function as a barrier against corneal neovascularization. However, the exact mechanisms regulating this remain unknown. In this study, the limbal epithelial stem cell (LESC) marker ABCB5 was used to investigate the role of LESCs in corneal neovascularization. In an ABCB5KO model, a mild but significant increase of limbal lymphatic and blood vascular network complexity was observed in developing mice (4 weeks) but not in adult mice. Conversely, when using a cornea suture model, the WT animals exhibited a mild but significant increase in the number of lymphatic vessel sprouts compared to the ABCB5KO, suggesting a contextual anti-lymphangiogenic effect of ABCB5 on the limbal vasculature during development, but a pro-lymphangiogenic effect under inflammatory challenge in adulthood. In addition, conditioned media from ABCB5-positive cultured human limbal epithelial cells (ABCB5+) stimulated human blood and lymphatic endothelial cell proliferation and migration. Finally, a proteomic analysis demonstrated ABCB5+ cells have a pro(lymph)angiogenic as well as an anti-inflammatory profile. These data suggest a novel dual, context-dependent role of ABCB5+ LESCs, inhibiting developmental but promoting inflammatory (lymph)angiogenesis in adulthood and exerting anti-inflammatory effects. These findings are of high clinical relevance in relation to LESC therapy against blindness.

Comparison of the Behavior of 3D-Printed Endothelial Cells in Different Bioinks.

Biomaterials with characteristics similar to extracellular matrix and with suitable bioprinting properties are essential for vascular tissue engineering. In search for suitable biomaterials, this study investigated the three hydrogels alginate/hyaluronic acid/gelatin (Alg/HA/Gel), pre-crosslinked alginate di-aldehyde with gelatin (ADA-GEL), and gelatin methacryloyl (GelMA) with respect to their mechanical properties and to the survival, migration, and proliferation of human umbilical vein endothelial cells (HUVECs). In addition, the behavior of HUVECs was compared with their behavior in Matrigel. For this purpose, HUVECs were mixed with the inks both as single cells and as cell spheroids and printed using extrusion-based bioprinting. Good printability with shape fidelity was determined for all inks. The rheological measurements demonstrated the gelling consistency of the inks and shear-thinning behavior. Different Young's moduli of the hydrogels were determined. However, all measured values where within the range defined in the literature, leading to migration and sprouting, as well as reconciling migration with adhesion. Cell survival and proliferation in ADA-GEL and GelMA hydrogels were demonstrated for 14 days. In the Alg/HA/Gel bioink, cell death occurred within 7 days for single cells. Sprouting and migration of the HUVEC spheroids were observed in ADA-GEL and GelMA. Similar behavior of the spheroids was seen in Matrigel. In contrast, the spheroids in the Alg/HA/Gel ink died over the time studied. It has been shown that Alg/HA/Gel does not provide a good environment for long-term survival of HUVECs. In conclusion, ADA-GEL and GelMA are promising inks for vascular tissue engineering.

Silencing of FGF6 hampers aerobic glycolysis and angiogenesis in bladder cancer by regulating PI3K/Akt and MAPK signaling pathways.

Metabolic abnormalities and uncontrolled angiogenesis are two vital features of malignant tumors. Although fibroblast growth factor 6 (FGF6) is known to promote the proliferation and migration of bladder cancer (BC) cells, its influences on aerobic glycolysis and angiogenesis in BC remain unclear. Gene expression at messenger RNA and protein levels were examined by reverse transcription-quantitative polymerase chain reaction and Western blot analyses, respectively. Lactate production and glucose uptake in BC cells were evaluated by performing aerobic glycolysis assays. A vasculogenic mimicry assay was executed for assessing the angiogenesis of BC cells. The viability, migration, and angiogenesis of human umbilical vein endothelial cells (HUVECs) cocultured with supernatants of BC cells were detected using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromideassay, wound healing assay, and tube formation assay.It was found that FGF6 displayed a high level in BC cell lines. Silencing of FGF6 reduced the levels of lactate production, glucose uptake, and the expression of angiogenic factors and glycolytic enzymes in BC cells, which also inhibited the viability and migration of HUVECs. In addition, FGF6 depletion or aerobic glycolysis inhibitor 2-deoxy-d-glucosetreatment decreased the total branching length and intersection number of both BC cells and HUVECs. Moreover, glucose or lactate treatment reversed FGF6-induced suppression of cell viability, migration, tube formation, and vasculogenic mimicry. The activation of the phosphatidylinositol-3-kinase(PI3K)/protein kinase B(Akt) and mitogen-activated protein kinase(MAPK) signaling pathways was blocked by silenced FGF6. Furthermore, PI3K/Akt inhibitor (LY2940002) and p38-MAPK inhibitor (SB203580) inhibited the levels of aerobic glycolysis-related proteins.In conclusion, FGF6 knockdown suppressed aerobic glycolysis, thereby inhibiting angiogenesis in BC via regulation of the PI3K/Akt and MAPK signaling pathways.

Mechanism of salidroside against coronary artery disease by network pharmacology analysis

BackgroundRosenroot (Rhodiola rosea) is a traditional Chinese herbal medicine. It has been used to treat patients with coronary artery disease (CAD). Salidroside is the main active constituent of rosenroot. This study was designed to explore the mechanism of salidroside in treating CAD and its role in angiogenesis in CAD systematically.MethodsIn this study, potential targets related to salidroside and CAD were obtained from public databases. Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), Disease Ontology (DO) and CellMarker enrichment analyses were performed. The binding of salidroside to angiogenesis-related targets was assessed by PyMOL and Ligplot. Furthermore, the effects of salidroside on collateral circulation were evaluated by correlation analysis of these angiogenesis-related targets with the coronary flow index (CFI), and the influence of salidroside on human umbilical vein endothelial cell (HUVEC) proliferation and migration was assessed.ResultsEighty-three targets intersected between targets of salidroside and CAD. GO and KEGG analyses indicated that salidroside mainly treated CAD through angiogenesis and anti-inflammatory action. There were 12 angiogenesis-related targets of salidroside in coronary heart disease, among which FGF1 (r = 0.237, P = 2.597E-3), KDR (r = 0.172, P = 3.007E-2) and HIF1A (r = -0.211, P = 7.437E-3) were correlated with the coronary flow index (CFI), and salidroside docked well with them. Finally, cell experiments confirmed that salidroside promoted the proliferation and migration of HUVECs.ConclusionsThis study revealed the potential molecular mechanism of salidroside on angiogenesis in CAD and provided new ideas for the clinical application of salidroside in the treatment of CAD.

AATF inhibition exerts antiangiogenic effects against human hepatocellular carcinoma.

Angiogenesis is a key factor in the growth and metastasis of hepatic tumors and thus a potential therapeutic target in hepatocellular carcinoma (HCC). In this study, we aim to identify the key role of apoptosis antagonizing transcription factor (AATF) in tumor angiogenesis and its underlying mechanisms in HCC. HCC tissues were analyzed for AATF expression by qRT-PCR and immunohistochemistry. Stable clones of control and AATF knockdown (KD) were established in human HCC cells. The effect of AATF inhibition on the angiogenic processes was determined by proliferation, invasion, migration, chick chorioallantoic membrane (CAM) assay, zymography, and immunoblotting techniques. We identified high levels of AATF in human HCC tissues compared to adjacent normal liver tissues, and the expression was found to be correlated with the stages and tumor grades of HCC. Inhibiting AATF in QGY-7703 cells resulted in higher levels of pigment epithelium-derived factor (PEDF) than controls due to decreased matric metalloproteinase activity. Conditioned media from AATF KD cells inhibited the proliferation, migration, and invasion of human umbilical vein endothelial cells as well as the vascularization of the chick chorioallantoic membrane. Furthermore, the VEGF-mediated downstream signaling pathway responsible for endothelial cell survival and vascular permeability, cell proliferation, and migration favoring angiogenesis was suppressed by AATF inhibition. Notably, PEDF inhibition effectively reversed the anti-angiogenic effect of AATF KD. Our study reports the first evidence that the therapeutic strategy based on the inhibition of AATF to disrupt tumor angiogenesis may serve as a promising approach for HCC treatment.

Jujuboside B suppresses angiogenesis and tumor growth via blocking VEGFR2 signaling pathway

Jujuboside B (JuB), one of the main active triterpenoid saponins from the traditional Chinese medicine Ziziphus jujuba, possesses a wide range of pharmacological activities. However, it is unknown whether JuB can inhibit tumor angiogenesis, a crucial step in solid tumor growth. In this study, we found that JuB significantly inhibited the proliferation, migration, and tube formation of human umbilical vein endothelial cells in a dose-dependent manner. JuB also suppressed angiogenesis in chick embryo chorioallantoic membranes and Matrigel plugs. Moreover, through angiogenesis inhibition, JuB delayed the growth of human HCT-15 colorectal cancer xenograft in mice. Western blot assay demonstrated that JuB inhibited the phosphorylation of VEGFR2 and its key downstream protein kinases, such as Akt, FAK, Src, and PLCγ1. In conclusion, the antiangiogenic potency and molecular mechanism of JuB are revealed for the first time, indicating that this triterpene saponin may be further explored as a potential drug candidate or lead compound for antiangiogenic cancer therapy.

Bioprinted Scaffold Remodels the Neuromodulatory Microenvironment for Enhancing Bone Regeneration

AbstractHerein, a 3D bioprinted scaffold is proposed, containing a calcitonin gene‐related peptide (CGRP) and the β‐adrenergic receptor blocker propranolol (PRN) as a new method to achieve effective repair of bone defects. By leveraging the neuromodulation mechanism of bone regeneration, CGRP and PRN loaded mesoporous silica nanoparticles are added into a hybrid bio‐ink, which initially contains gelatin methacrylate, Poly (ethylene glycol) diacrylate and bone marrow mesenchymal stem cells (BMSCs). Subsequently, the optimized bio‐ink is used for 3D bioprinting to create a composite scaffold with a pre‐designed micro‐nano hierarchical structure. The migration and tube formation of human umbilical vein endothelial cells (HUVECs) can be promoted by the scaffold, which is beneficial to the formation of a new capillary network during the bone repair process. With the release of CGRP from the scaffold, the secretion of neuropeptides by sensory nerves is simulated. Meanwhile, the release of PRN can inhibit the binding process of catecholamine to β‐adrenergic receptor, co‐promoting the osteogenic differentiation of BMSCs with CGRP and silicon ions, which will effectively enhance bone repair of a critical‐sized cranial defect in a rat model. In conclusion, this study provides a promising strategy for bone defect repair by understanding the neuromodulatory mechanisms during bone regeneration.