Angiogenic Potential Of Endothelial Cells Research Articles

Omentum Extracellular Matrix-Silk Fibroin Hydroscaffold Promotes Wound Healing through Vascularization and Tissue Remodeling in the Diabetic Rat Model.

Nonhealing diabetic wounds are often associated with significant mortality and cause economic and clinical burdens to the healthcare system. Herein, a biomimetic hydroscaffold is developed using omentum tissue-derived decellularized-extracellular matrix (dECM) and silk fibroin (SF) proteins that associate the behavior of a collagenous fibrous scaffold and a hydrogel to reproduce all aspects of the provisional skin tissue matrix. The chemical cross-linker-free in situ gelation property of the two types of SF proteins from Bombyx mori and Antheraea assamensis ensures the adherence of dECM with surrounding tissue on the wound bed, circumventing further suturing. The physicochemical and mechanical properties of the composite hydroscaffold (SF-dECM) were thoroughly evaluated. The hydroscaffolds were found to support the growth and proliferation of human dermal fibroblasts and influence the angiogenic potential of endothelial cells under in vitro conditions. Furthermore, the healing efficacy of the composites was evaluated by generating full-thickness wounds on a streptozotocin-induced diabetic rat model. The presence of dECM components in the composite facilitated the rate of wound closure, granulation tissue formation, and re-epithelialization by providing intrinsic cues to advance the inflammatory stage and stimulating angiogenesis. Collectively, as an off-the-shelf wound dressing requiring only a single topical administration, the SF-dECM hydroscaffold is a promising, cost-effective dressing for the management of chronic diabetic wounds.

Angiogenesis is promoted by hypoxic cervical carcinoma-derived extracellular vesicles depending on the endothelial cell environment

IntroductionCancer needs perfusion for its growth and metastasis. Cancer cell-derived extracellular vesicles (CA-EVs) alter the tumor microenvironment (TME), potentially promoting angiogenesis. We hypothesize that conditions in the tumor, e.g., hypoxia, and in the target cells of the TME, e.g., nutrient deprivation or extracellular matrix, can affect the angiogenic potential of CA-EVs, which would contribute to explaining the regulation of tumor vascularization and its influence on cancer growth and metastasis. MethodsCA-EVs were isolated and characterized from cervical carcinoma cell lines HeLa and SiHa cultured under normoxia and hypoxia, and their angiogenic potential was evaluated in vitro in three endothelial cells (ECs) lines and aortic rings, cultured in basal (growth factor-reduced) or complete medium. ResultsHypoxia increased EV production 10–100 times and protein content 2–4 times compared to normoxic CA-EVs. HeLa-EVs contained six times more RNA than SiHa-EVs, and this concentration was not affected by hypoxia. Treatment with CA-EVs increased tube formation and sprouting in ECs and aortic rings cultured in basal medium and long-term stabilized the stablished vascular networks formed by ECs cultured in complete medium. ConclusionHypoxia differentially affects CA-EVs in a cell line-dependent manner. The cellular environment (nutrient availability and extracellular matrix scaffold) influences the effect of CA-EV on the angiogenic potential of ECs.

Cobalt-doped layered hydroxide coating on titanium implants promotes vascularization and osteogenesis for accelerated fracture healing

Angiogenesis at the fracture site plays crucial roles in the endogenous osteogenesis process and is a prerequisite for the efficient repair of implant fixed bone defects. To improve the peri-implant vascularization of titanium implant for accelerating defect healing, we developed a Co-doped Mg–Al layered hydroxide coating on the surface of titanium using hydrothermal reaction and then modified the surface with gallic acid (Ti-LDH/GA). Gallic acid coating enabled the sustained release of Co2+ and Mg2+ to the defect site over a month. Ti-LDH/GA treatment profoundly stimulated the angiogenic potential of endothelial cells by upregulating the vascularization regulators such as vascular endothelial growth factor VEGF) and hypoxia-inducible factor-1α (HIF-1α), leading to enhanced osteogenic capability of mesenchymal stem cells (MSCs). These pro-bone healing benefits were attributed to the synergistic effects of Co ions and Mg ions in promoting angiogenesis and new bone formation. These insights collectively suggested the potent pro-osteogenic effect of Ti-LDH/GA through leveraging peri-implant vascularization, offering a new approach for developing biofunctional titanium implants.

Interaction between Macrophages and Adipose Stromal Cells Increases the Angiogenic and Proliferative Potential of Pregnancy-Associated Breast Cancers.

Pregnancy associated breast cancers (PABCs) exhibit increased aggressiveness and overall poorer survival. During lactation, changes take place in the breast tissue microenvironment that lead to increased macrophage recruitment and alterations in adipose stromal cells (ASC-Ls). The interaction of these cells in PABCs could play a role in the increased aggressiveness of these cancers. We utilized an in vitro co-culture model to recreate the interactions of ASC-Ls and macrophages in vivo. We performed qRT-PCR to observe changes in gene expression and cytokine arrays to identify transcriptional changes that result in an altered microenvironment. Additionally, functional assays were performed to further elicit how these changes affect tumorigenesis. The co-culture of ASC-Ls and macrophages altered both mRNA expression and cytokine secretion in a tumor promoting manner. Tumorigenic cytokines, such as IL-6, CXCL1, CXCL5, and MMP-9 secretion levels, were enhanced in the co-culture. Additionally, conditioned media from the co-culture elevated the tumor cell proliferation and angiogenic potential of endothelial cells. These finds indicate that the changes seen in the microenvironment of PABC, specifically the secretion of cytokines, play a role in the increased tumorigenesis of PABCs by altering the microenvironment to become more favorable to tumor progression.

Abstract P2077: Caspase8ap2 M6a Mrna Methylation Enhances Cardiomyocyte Apoptosis Following Myocardial Ischemia

Objective: Heart failure is the leading cause of morbidity and mortality in the United States. Cardiomyocyte death is one of the critical mediators of cardiac dysfunction following ischemic injury. However, the role of posttranscriptional regulation on cell-death-associated genes is not well explored during ischemic heart failure. N6-methyladenosine (m6A) is the most abundant and conserved chemical modification found in eukaryotic mRNA and is associated with mRNA synthesis and metabolism. We have recently shown that m6A RNA methylation regulates the angiogenic potential of endothelial cells following ischemic injury. However, the role of m6A mRNA methylation in cardiac cell death following ischemic injury is not well defined. Methods: Cardiac function (using echocardiography) was measured at baseline and four weeks post-MI in mice. In the end, mice were euthanized, and hearts were excised for biochemical and histological analysis. Cardiac cell death was assessed in H9C2 cells and cardiomyocytes using FACs (Annexin V and PI) and TUNNEL assay. The Casp8AP2 stability were measured using actinomycin D assay. To demonstrate the molecular mechanism, METTL3 was inhibited using siRNA or shRNA. Results: The cardiac function was significantly reduced following MI. Enhanced m6A mRNA methylation was observed in mice hearts following MI. Histological (TUNEL assay) and biochemical (WB: Caspase 8, cleaved cas8, caspase3, cleaved caspase3, Casp8AP2) analysis showed increased myocytes apoptosis in mice following MI. At the molecular level, hypoxic stress (nutrient depiviation+1%O2) induces global m6A mRNA methylation, which leads to Cas8AP2 gene stability and activation of extrinsic apoptotic pathways in cardiomyocytes in vitro. In contrast, METTL3 inhibition significantly reduced Cas8AP2 mRNA methylation, enhanced its degradation, and ultimately reduced cardiomyocyte cell death. Conclusion: In conclusion, we have shown that increased Casp8AP2 m6A RNA methylation enhances its stability and activates caspase-3 cleavage, which eventually leads to cardiomyocyte death following myocardial ischemia. The reduction of the RNA methylation through METTL3 inhibition (methyltransferase) reduced hypoxia-induced apoptosis of cardiomyocytes.

Enhanced expression of pentraxin-3 in glioblastoma cells correlates with increased invasion and IL8-VEGF signaling axis

Glioblastoma (GB) is highly invasive and resistant to multimodal treatment partly due to distorted vasculature and exacerbated inflammation. The aggressiveness of brain tumors may be attributed to the dysregulated release of angiogenic and inflammatory factors. The glycoprotein pentraxin-3 (PTX3) is correlated with the severity of some cancers. However, the mechanism responsible for the invasive oncogenic role of PTX3 in GB malignancy remains unclear. In this study, we examined the role of PTX3 in GB growth, angiogenesis, and invasion using in vitro and in vivo GB models, proteomic profiling, molecular and biochemical approaches. Under in vitro conditions, PTX3 over-expression in U87 cells correlated with cell cycle progression, increased migratory potential, and proliferation under hypoxic conditions. Conditioned media containing PTX3 enhanced the angiogenic potential of endothelial cells. While silencing of PTX3 by siRNA decreased the proliferation, migration, and angiogenic potential of U87 cells in vitro. Importantly, PTX3 over-expression increased tumor growth, angiogenesis, and invasion in an orthotopic mouse model. Higher levels of PTX3 in these tumors were associated with the upregulation of inflammatory and angiogenic markers including interleukin-8 (IL-8) and vascular endothelial growth factor (VEGF), but decreased levels of thrombospondin-1, an anti-angiogenic factor. Mechanistically, exogenous production of PTX3 triggered an IKK/NFκB signaling pathway that enhances the expression of the motility genes AHGEF7 and Rac1. Taken together, PTX3 expression is dysregulated in GB. PTX3 may augment invasion through enhanced angiogenesis in the GB microenvironment through the IL8-VEGF axis. Thus, PTX3 may represent a potential therapeutic target to mitigate the aggressive behavior of gliomas.

An improved osseointegration of metal implants by pitavastatin loaded multilayer films with osteogenic and angiogenic properties

An increasing number of works have highlighted the importance of metal implants surface modification in enhancing bone defect healing through the synergistic osteogenesis-angiogenesis regulation. Studies have shown that pitavastatin has the effect of promoting osteogenesis and angiogenesis. However, how to prepare pitavastatin functionalized implants and how pitavastatin regulates the synergies of osteogenesis and angiogenesis around implants as well as the related mechanisms remain unclear. In the present study, multilayer films with osteogenic and angiogenic properties were constructed on pure titanium substrates via the layer-by-layer assembly of pitavastatin-loaded β-cyclodextrin grafted chitosan and gelatin. In vitro experiments demonstrated that locally applied pitavastatin could dramatically enhance osteogenic potential of mesenchymal stem cells (MSCs) and angiogenic potential of endothelial cells (ECs). Moreover, pitavastatin loaded multilayer films could regulate the paracrine signaling mediated crosstalk between MSCs and ECs, and indirectly increase the angiogenic potential of MSCs and osteogenic potential of ECs via multiple paracrine signaling. The results of subcutaneous and femur implantation confirmed that locally released pitavastatin had potentially triggered a chain of biological events: mobilizing endogenous stem cells and ECs to the implant-bone interface, in turn facilitating coupled osteogenesis and angiogenesis, and eventually enhancing peri-implant osseointegration. This study enlarges the application scope of pitavastatin and provides an optional choice for developing a multifunctional bioactive coating on the surfaces of mental implants.

A dual-ink 3D printing strategy to engineer pre-vascularized bone scaffolds in-vitro

A functional vascular supply is a key component of any large-scale tissue, providing support for the metabolic needs of tissue-remodeling cells. Although well-studied strategies exist to fabricate biomimetic scaffolds for bone regeneration, success rates for regeneration in larger defects can be improved by engineering microvascular capillaries within the scaffolds to enhance oxygen and nutrient supply to the core of the engineered tissue as it grows. Even though the role of calcium and phosphate has been well understood to enhance osteogenesis, it remains unclear whether calcium and phosphate may have a detrimental effect on the vasculogenic and angiogenic potential of endothelial cells cultured on 3D printed bone scaffolds. In this study, we presented a novel dual-ink bioprinting method to create vasculature interwoven inside CaP bone constructs. In this method, strands of a CaP ink and a sacrificial template material was used to form scaffolds containing CaP fibers and microchannels seeded with vascular endothelial and mesenchymal stem cells (MSCs) within a photo-crosslinkable gelatin methacryloyl (GelMA) hydrogel material. Our results show similar morphology of growing vessels in the presence of CaP bioink, and no significant difference in endothelial cell sprouting was found. Furthermore, our initial results showed the differentiation of hMSCs into pericytes in the presence of CaP ink. These results indicate the feasibility of creating vascularized bone scaffolds, which can be used for enhancing vascular formation in the core of bone scaffolds.

HIF-1 stabilization exerts anticancer effects in breast cancer cells in vitro and in vivo

Tumor hypoxia and high activity of hypoxia-inducible factor-1 (HIF-1) correlate with adverse disease outcomes, malignancy, resistance to therapy and metastasis. Nonetheless, recent studies indicate that under certain circumstances, HIF-1 stabilization may exert protective effects and even decrease tumor cell aggressiveness.This study aimed to characterize the potential anticancer effect of molidustat (BAY 85-3934), the prolyl hydroxylase (PHD) inhibitor and HIF-1 stabilizator. We confirmed that molidustat stabilizes HIF-1α and induces the expression of vascular endothelial growth factor (VEGF) in MDA-MB-231 breast cancer cells, to a similar or even greater extent than hypoxia. Interestingly, decreased cell survival and colony formation capabilities, together with S/G2 cell cycle arrest, were observed after treatment with PHD inhibitor. Importantly, molidustat enhanced the effectiveness of the chemotherapeutic drug, gemcitabine, on cancer cells. Finally, the xenograft model revealed decreased tumor growth in vivo after molidustat treatment. Both in vitro and in vivo analysis showed no differences in the angiogenic potential of endothelial cells treated with tumor-conditioned media or vascularization of the MDA-MB-231 xenografts, respectively.In summary, molidustat treatment exhibits an inhibitory effect on breast cancer cell survival, self-renewal capacity and potentiates the efficacy of chemotherapeutic gemcitabine.

Leptin-Induced Angiogenesis of EA.Hy926 Endothelial Cells via the Akt and Wnt Signaling Pathways In Vitro and In Vivo.

Angiogenesis involves the activation of endothelial cells followed by capillary formation. Leptin, the protein product of the ob gene, can induce the angiogenic potential of endothelial cells. However, the underlying cellular mechanism still remains to be elicited. We firstly evaluated the in vitro effects of leptin on proliferation and angiogenic differentiation of endothelial cell line EA.hy926. Leptin was found to potently induced cell proliferation, expression of angiogenic gene, migration and tube formation. Then we investigated the roles of the Akt and Wnt signaling pathways in the aforementioned processes. It showed that Akt and Wnt signaling pathways could be activated by leptin, while inhibition of the Akt and Wnt signaling pathways by siRNAs effectively blocked the leptin-induced angiogenesis. Finally, we used electrospinning to fabricated leptin-immobilized linear poly(L-lactide-co-caprolactone) (PLCL)-leptin. The in vivo vessel formation of PLCL-leptin was evaluated using subcutaneous implants in Sprague-Dawley rats. The histological and immunofluorescence revealed that cell infiltration with PLCL-leptin was much more significant than that with the control PLCL group. More importantly, the number of laminin+ vessels and CD31+ cells in PLCL-leptin grafts was significantly higher than in control grafts. The study demonstrated that it is via Akt and Wnt signaling pathways that leptin promotes the proliferation and angiogenic differentiation of endothelial cells and the capacity of endogenous tissue regeneration makes the novel leptin-conjugated PLCL promising materials for grafts.

Metformin Effect on Endocan Biogenesis in Human Endothelial Cells Under Diabetic Condition

Endocan is a novel endothelium-derived proteoglycan and may play a role in endothelial cells activity under diabetic conditions. Here, we evaluated the effect of high glucose concentration (30mmol glucose) on endocan level in presence or absence of metformin in human umbilical vein endothelial cells (HUVECs). Cells were incubated with 30mmol glucose for 72h. High glucose content, metformin (2.5 to 500mmol) and compound C (10mmol) effects were assessed on cell viability. HUVECs migration was studied by scratch test. The changes in endocan expression and protein level were evaluated by RT-PCR, ELISA and flow cytometry assays. Griess reaction was used to measure NO levels. Functional activity of endothelial cells was monitored related to lipoprotein lipase activity using Dil-Ac-LDL uptake. p-AMPK/AMPK ratio was assessed by western blotting. Cells viability significantly was reduced under high glucose condition (p<0.05). 30mmol glucose inhibited HUVECs migration, whereas these features were improved by 50mmol metformin (p<0.05). Endocan transcription and protein levels were increased in diabetic HUVECs exposed to metformin (p<0.05). Metformin increased NO production in HUVECs under high glucose condition (p<0.001). Metformin increased LDL uptake capacity under high glucose condition (p<0.05). The addition of compound C blunted these effects. Western blot analysis confirmed the increase of p-AMPK/AMPK ratio in metformin-treated cells. Data demonstrated that metformin could promote angiogenic potential of endothelial cells which its reduction is a main cause in the development of diabetic foot ulcer, probably by the regulation of endocan dynamics under high glucose condition.

SAT-392-Dickkopf-1 from hepatocellular carcinoma cells promotes the angiogenic potential of endothelial cells by activating the VEGFR-2 signaling pathway

SAT-392-Dickkopf-1 from hepatocellular carcinoma cells promotes the angiogenic potential of endothelial cells by activating the VEGFR-2 signaling pathway

Endothelial cells from pulmonary endarterectomy specimens possess a high angiogenic potential and express high levels of hepatocyte growth factor

BackgroundImpaired angiogenesis is assumed to be an important factor in the development of chronic thromboembolic pulmonary hypertension (CTEPH). However, the role of endothelial cells (ECs) in CTEPH remains unclear. The aim of this study was to investigate the angiogenic potential of ECs from pulmonary endarterectomy (PEA) specimens.MethodsWe isolated ECs from PEA specimens (CTEPH-ECs) and control EC lines from the intact pulmonary arteries of patients with peripheral lung cancers, using a MACS system. These cells were analyzed in vitro including PCR-array analysis, and the PEA specimens were analyzed with immunohistochemistry. Additionally, the serum HGF levels were determined in CTEPH patients.ResultsA three-dimensional culture assay revealed that CTEPH-ECs were highly angiogenic. An angiogenesis-focused gene PCR array revealed a high expression of hepatocyte growth factor (HGF) in CTEPH-ECs. The high expression of HGF was also confirmed in the supernatant extracted from PEA specimens. The immunohistochemical analysis showed expression of HGF on the surface of the thrombus vessels. The serum HGF levels in CTEPH patients were higher than those in pulmonary thromboembolism survivors.ConclusionOur study suggests that there are ECs with pro-angiogenetic character and high expression of HGF in PEA specimens. It remains unknown how these results are attributable to the etiology. However, further investigation focused on the HGF pathway may provide novel diagnostic and therapeutic tools for patients with CTEPH.

Role of Notch1 in the arterial specification and angiogenic potential of mouse embryonic stem cell-derived endothelial cells

BackgroundEndothelial cells have been shown to mediate angiogenesis in ischemic injury sites and contribute to the repair of damaged tissues. However, the treatment of ischemic disease requires a significant number of endothelial cells, which are difficult to isolate from patients. Embryonic stem cells have been considered a potential source of therapeutic cells due to their unlimited self-renewal and pluripotent properties. With regard to vascular development, Notch1 has been established as a key regulator of the specification of arterial endothelial cells.MethodsUsing a doxycycline-induced expression system of the intracellular domain of Notch1, we explored the role of Notch1 in the differentiation of embryonic stem cells to arterial endothelial cells. The therapeutic effect of the arterial endothelial cells was investigated in a murine hindlimb ischemia model. The blood perfusion rate in the ischemic limb was determined by laser Doppler perfusion imaging, and vasculogenesis was quantified using immunocytochemistry.ResultsInduced expression of the intracellular domain of Notch1 increased the levels of endothelial markers, such as CD31 and VE-cadherin, in differentiated endothelial cells. Induction of intracellular domain of Notch1 stimulated expression of the arterial-type endothelial cell markers (Nrp1 and Ephrin B2), but not the venous-type endothelial cell markers (Nrp2 and Coup-TFII). In addition, overexpression of intracellular domain of Notch1 resulted in increased expression of CXCR4, a chemokine receptor involved in vascular development. Induction of intracellular domain of Notch1 increased endothelial tube formation and migration of differentiated endothelial cells. Intramuscular administration of Notch1-induced arterial endothelial cells was more effective than administration of the control endothelial cells in restoring the blood flow in an ischemic hindlimb mouse model. Transplantation of Notch1-induced arterial endothelial cells augmented the number of blood vessels and incorporation of endothelial cells into newly formed blood vessels.ConclusionsThese results suggest that Notch1 promotes endothelial maturation and arterial specification during the differentiation of embryonic stem cells to endothelial cells and increases the angiogenic potential of endothelial cells.

Effects of preadipocytes derived from mice fed with high fat diet on the angiogenic potential of endothelial cells

Background and aimsObesity promotes a persistent inflammatory process in the adipose tissue, activating the endothelium and leading to vascular dysfunction. Preadipocytes can interact with endothelial cells in a paracrine way stimulating angiogenesis. However, the potential of preadipocytes from adipose tissue of high fat diet (HFD) fed animal to stimulate angiogenesis has not been evaluated yet. The aim of this study was to investigate the effects of such diet on the angiogenic potential of preadipocytes in a mice model. Methods and resultsWe have evaluated body weight gain, fasting glucose levels and insulin resistance, mRNA expression in preadipocytes and endothelial cells after co-culture with preadipocytes, in vivo vascular function and in vitro endothelial cell migration and tubulogenesis. High fat diet promoted an increase in body weight, glycemic index and insulin resistance in mice. Preadipocytes mRNA expression of factors involved in angiogenesis was higher in these animals. In endothelial tEnd cells mRNA expression of factors involved in vessel growth were higher after co-culture with preadipocytes derived from mice fed with HFD. Although no significant differences were observed in in vivo vasodilatation response between control and HFD groups, endothelial tEnd cells showed an increase in migration and tubulogenesis when cultivated with conditioned media from preadipocytes derived from mice fed with HFD. ConclusionHypoxic and growth factors produced by preadipocytes derived from mice fed with HFD have higher capacity than preadipocytes derived from mice fed with standard diet to stimulate the angiogenic potential of endothelial cells, contributing to vascular disorders in obesity.

Metformin prevents nerve growth factor-dependent proliferative and proangiogenic effects in epithelial ovarian cancer cells and endothelial cells.

Background:Epithelial ovarian cancer (EOC) is characterized by exacerbated angiogenesis regulated by proangiogenic and growth factors. Nerve growth factor (NGF) is overexpressed in EOC where it promotes proliferation as well as survival and is considered a proangiogenic factor. Metformin, a drug commonly used in the treatment of diabetes, is attributed to antineoplastic effects, but the underlying mechanisms remain unknown. Given that current therapies yield modest results in EOC patients, the aim of this study was to determine the effects of metformin on NGF-enhanced proliferation of EOC cells and the angiogenic potential of endothelial cells.Methods:A2780 (EOC), HOSE (human ovarian surface epithelial) and EA.hy926 (endothelial) cells were treated with NGF and metformin. Cell viability, cell proliferation and cell cycle were evaluated in all three cell lines, and the angiogenic potential in endothelial EA.hy926 cells.Results:NGF enhanced cell proliferation in A2780, HOSE and EA.hy926 cells (p < 0.05), while metformin treatment decreased cell proliferation in A2780 and EA.hy926 cells (p < 0.05). Moreover, the NGF-enhanced angiogenic score in EA.hy926 cells was prevented by metformin (p < 0.05).Conclusions:Given that NGF plays a significant role in EOC progression, our current findings suggest that metformin holds considerable promise as an adjuvant treatment in ovarian cancer.

PROFILING AND CHARACTERIZING MICRORNAS IN PATIENTS WITH HEREDITARY HEMORRHAGIC TELANGIECTASIA

Hereditary hemorrhagic telangiectasia (HHT) is a rare, autosomal dominant, vascular disorder that affects 1 in 5000 to 8000 individuals worldwide. HHT is characterized by vascular abnormalities, such as telangiectasias and arteriovenous malformations (AVMs). Telangiectasias are dilated skin and mucocutaneous blood vessels that are susceptible to spontaneous hemorrhage. AVMs are direct connections between arteries and veins, without intervening capillaries, and can cause significant right to left shunts. HHT arises from heterozygous mutations in at least 3 genes, ENG, ACVRL1, and SMAD4. Haploinsufficiency of their respective protein products perturb TGF signalling, affecting the angiogenic potential of endothelial cells (ECs) and the migratory capacity of monocytes, contributing to vascular malformations in HHT. MicroRNAs (miRs) are small non-coding RNAs that post-transcriptionally target mRNA transcripts. We aimed to profile and characterize miRs in HHT patient derived early endothelial progenitor cells (eEPCs) and peripheral blood mononuclear cells (PBMCs) to explore their potential role in HHT pathogenesis. EEPCs are early outgrowth ECs cultured from the mononuclear cell layer (buffy coat) of peripheral blood. They have been shown to augment angiogenesis in a paracrine manner by secreting various growth factors and are a widely accepted surrogate for studying EC function 28 HHT patients with confirmed mutations, and 25 controls were recruited. 40 ml of peripheral blood was collected for eEPC culture and PBMC isolation. Total RNA was isolated from PBMCs and eEPCs. PBMC and eEPC expression profiling was conducted with a human miR array analysis. Dysregulated miRs identified from the analyses were then validated with RT-qPCR. Significant differences were determined using a two-tailed t-test. Of the 800 miRs screened, 167 and 121 dysregulated miRs were identified in PBMCs and eEPCs, respectively. Selected PBMC miRs (MiRs-28-5p, -30b-5p, -361-3p and -374a-5p) and eEPC miRs (miRs-132-3p, -221-3p and -424-5p) were validated with RT-qPCR. PBMC miR-361-3p (p=0.025) known to target IGF1, and eEPC miR-132-3p (p=0.047) known to target SMAD2, were found to be significantly decreased compared to controls. Subsequently, IGF1 messenger RNA (mRNA) levels were significantly increased (p=0.005) in PBMCs of HHT patients compared to controls. Our results show dysregulation of select miRs in PBMCs and eEPCs from HHT patients. The finding of increased IGF1 mRNA and its possible relation to decreased miR-361-3p levels is both novel and exciting. This may represent a putative pathogenic mechanism involved in HHT, and may provide a unique miR therapeutic target for the treatment of clinical manifestations. National Institutes of Health (NIH)

Effect of Cytokines on the Formation Tube-Like Structures by Endothelial Cells in the Presence of Trophoblast Cells.

Despite ample data on cytokine secretion in the uteroplacental interface, the influence of microenvironment cells, in particular, trophoblast cells on angiogenesis and the role of cytokines in this process remain poorly studied. We studied the influence of cytokines on the formation of tube-like structures by endothelial cells in the presence of trophoblast cells and showed that trophoblast cells suppressed the angiogenic potential of endothelial cells. Antiangiogenic cytokines IFN-γ, IL-10, TNF-α, and TGFβ via modulation of trophoblast cells stimulated the formation of tube-like structures by endothelial cells. In the co-culture of endothelial and trophoblast cells, the effects of cytokines changed and they gained additional regulatory functions.

The association between interleukin-19 concentration and diabetic nephropathy

BackgroundInterleukin-19 (IL-19) is a newly discovered cytokine belonging to the Interleukin-10(IL-10) family. IL-19 have indispensable functions in many inflammatory processes and also can induce the angiogenic potential of endothelial cells. The purpose of present study was to investigate the relation of serum interleukin-19 (IL-19) levels with diabetic nephropathy (DN).MethodsTwo hundred study groups of patients with type 2 diabetes mellitus (T2DM) (109 males and 91 females) were recruited, included normoalbuminuria(n = 102), microalbuminuria(n = 72) and macroalbuminuria(n = 26) . The 50 healthy blood donors were enrolled for the control group. All subjects were assessed for: IL-19, High-sensitivity C-reactive protein (Hs-CRP), Cystatin C, urinary albumin excretion rate (UAE) and glycosylated hemoglobin A1c(HbA1c).ResultsThe serum IL-19 levels in DN patients were found to be significantly higher compared to controls. IL-19 levels were significantly positively correlated with Hs-CRP, Cystatin C, UAE and HbA1c(r = 0.623, 0.611,0.591 and 0.526 respectively, P < 0.01). Multivariable logistic regression analysis showed IL-19 levels (P = 0.01) were found to be independently associated with patients with DN.ConclusionsIL-19 is significantly positive correlated with UAE and Cystatin C. IL-19 may play an important role that contributes to the progression of diabetic nephropathy.

Activated thrombin-activatable fibrinolysis inhibitor attenuates the angiogenic potential of endothelial cells: potential relevance to the breast tumour microenvironment

Thrombin-activatable fibrinolysis inhibitor (TAFI) is a basic carboxypeptidase zymogen present in blood plasma. Proteolytic activation of TAFI by thrombin, thrombin in complex with the endothelial cell cofactor thrombomodulin, or plasmin results in an enzyme (TAFIa) that removes carboxyl-terminal lysine residues from protein and peptide substrates, including cell-surface plasminogen receptors. TAFIa is therefore capable of inhibiting plasminogen activation in the pericellular milieu. Since plasminogen activation has been linked to angiogenesis, TAFIa could therefore have anti-angiogenic properties, and indeed TAFIa has been shown to inhibit endothelial tube formation in a fibrin matrix. In this study, the TAFI pathway was manipulated by providing exogenous TAFI or TAFIa or by adding a potent and specific inhibitor of TAFIa. We found that TAFIa elicited a series of anti-angiogenic responses by endothelial cells, including decreased endothelial cell proliferation, cell invasion, cell migration, tube formation, and collagen degradation. Moreover, TAFIa decreased tube formation and proteolysis in endothelial cell culture grown alone and in co-culture with breast cancer cell lines. In accordance with these findings, inhibition of TAFIa increased secretion of matrix metalloprotease proenzymes by endothelial and breast cancer cells. Finally, treatment of endothelial cells with TAFIa significantly inhibited plasminogen activation. Taken together our results suggest a novel role for TAFI in inhibiting tumour angiogenic behaviors in breast cancer.